62 entries found for : sfn2001
Truitt WA, Stanton LA, Coolen LM (2001) Targeted lesions of Substance P receptor cells in L3-4 lumbar spinal cord severely impair male ejaculatory behavior. Neuroscience 2001 Abstracts 958.18. Society for Neuroscience, San Diego, CA.
Summary: Previously we demonstrated the existence of a spinothalamic pathway in the male rat where neural activation is specifically induced by ejaculation. This pathway consists of the parvocellular subparafascicular thalamic nucleus (SPFp) and the neurons projecting to the SPFp that are located in the lumbar spinal cord, specifically in laminae 10 and 7 of segments L3-L4. This neuron population coexpresses galanin (Gal) and substance P receptors (SPR). To test the hypothesis that these cells relay sensory information related to ejaculation, these neurons were lesioned by targeting the SPR with the neurotoxin SSP-saporin, a high affinity analogue of substance P conjugated to the ribosome inhibitor saporin. SSP-saporin (4 ng/µl) or an equal concentration of unconjugated saporin was injected bilaterally into L3-L4 region in either sexually experienced or naïve male Sprague Dawley rats (n=5 each). Sexual behavior was tested weekly, starting ten days following surgery. Following the sixth behavioral test, rats were sacrificed and spinal cord tissue was immunostained for GAL, SPR, or NeuN to assess specific and nonspecific tissue damage. In rats injected with unconjugated saporin, the Gal/SPR cell population was intact and sexual behavior was not altered. In contrast, in rats receiving SSP-saporin, Gal/SPR cell population was significantly decreased (77%) or absent. Moreover, SSP-saporin lesions severely disrupted ejaculatory behavior. Interestingly, display of mounts and intromissions remained intact. These results suggest that Gal/SPR cells in laminae 10 and 7 of L3-4 lumbar spinal cord may be essential for male ejaculatory behavior.
Related Products: SSP-SAP (Cat. #IT-11)
Dinh TT, Sanders NM, Pedrow C, Ritter S (2001) Selective immunotoxin lesion of spinally projecting norepineprhine and epinephrine (NE/E) neurons impairs the glucagon response to 2-deoxy-d-glucose (2DG). Neuroscience 2001 Abstracts 947.1. Society for Neuroscience, San Diego, CA.
Summary: Previous work has shown that the targeted immunotoxin, anti-dopamine ß-hydroxylase conjugated to saporin (DSAP), can be used to selectively destroy subpopulations of hindbrain NE/E neurons projecting to or through a particular DSAP injection site. Using this approach, we have shown that NE/E neurons projecting to the hypothalamus are required for feeding and glucocorticoid responses to 2DG-induced glucoprivation and those projecting spinally are required for the adrenal medullary response. In this study, we injected DSAP or unconjugated saporin (SAP) control solution into the spinal cord at T2-T4 to investigate the role of the spinally projecting NE/E neurons in glucagon secretion induced by 2DG (250 mg/kg). Controls injected spinally with unconjugated saporin (SAP) had a glucagon response that peaked at 308% of pre-2DG levels, while DSAP-injected rats had a significantly blunted response to 2DG, peaking at 197% of pre-2DG levels. 2DG-induced hyperglycemia also was impaired in the DSAP rats, but not in the SAP rats. Both SAP and DSAP rats had normal feeding and glucocorticoid responses to 2DG. Results suggest that spinally-projecting NE/E neurons participate in the neural control of glucagon secretion under conditions of glucose deficit. In combination with other findings, these results indicate that hindbrain NE/E neurons contribute to four major glucoregulatory responses (increased feeding, and increased secretion of glucagon, glucocorticoids and adrenal medullary epinephrine) through their projections to hypothalamic or spinal cord effector sites.
Related Products: Anti-DBH-SAP (Cat. #IT-03)
Fraley GS, Dinh TT, Ritter S (2001) Immunotoxin lesion of catecholaminergic neurons innervating the medial hypothalamus elevates basal expression of and attenuates glucoprivaation-induced increases in agouti gene related protein (AGRP) mRNA. Neuroscience 2001 Abstracts 947.2. Society for Neuroscience, San Diego, CA.
Summary: Catecholaminergic (CA) innervation of medial hypothalamic structures is necessary for glucoprivation-induced feeding, glucocorticoid secretion and Fos expression in the paraventricular (PVH) and arcuate nuclei of the hypothalamus. In this experiment, we tested the hypothesis that the 2-deoxy-D-glucose (2DG)-induced an increase in AGRP mRNA expression (reported recently by Sergeyev et al., 2000) also requires NE/E neurons. CA neurons innervating the medial hypothalamus were lesioned using the toxin, saporin, targeted for selective entry into NE/E neurons by conjugation with a monoclonal antibody against dopamine beta hydroxylase. This toxin (DSAP), or unconjugated saporin (SAP) control solution, was bilaterally microinjected into the PVH. DSAP rats with confirmed 2DG-induced feeding deficits (DSAP 1.7 +/- 0.29 g; SAP 5.1 +/- 0.31 g; p < 0.05) and controls were injected with 2DG (250 mg/kg), or saline and maintained for 2 hrs without food. Hypothalami were harvested and subjected to Northern blot analysis of AGRP mRNA. Blot analysis revealed that 2DG increased mRNA expression in SAP controls (2DG: 1.0 +/- 0.05 RDU; saline: 0.7 +/- 0.02, p< .05), but not in DSAP lesioned rats (2DG: 1.1 +/- 0.04; saline: 1.0 +/- 0.03). In addition, basal AGRP mRNA expression was significantly elevated in DSAP-lesioned rats compared to SAP controls (p < .05). These data suggest that basal AGRP gene expression is controlled by hindbrain CA neurons and that increased AGRP gene expression induced by glucoprivation also requires these neurons.
Related Products: Anti-DBH-SAP (Cat. #IT-03)
Ritter S, Dinh TT, Sanders NM, Pedrow C (2001) Selective immunotoxin lesion of hypothalamically-projecting norepinephrine/epinephrine (NE/E) neurons impairs the glucocorticoid response to glucoprivation. Neuroscience 2001 Abstracts 947.3. Society for Neuroscience, San Diego, CA.
Summary: Decreased glucose utilization triggers behavioral and neuroendocrine responses that increase blood glucose concentrations and delivery of glucose to the brain. These include stimulation of food intake and increased secretion of glucagon, adrenal E and glucocorticoids. In previous work utilizing the targeted immunotoxin, DSAP (saporin conjugated to a monoclonal antibody against dopamine β-hydroxylase), we demonstrated that hindbrain NE/E neurons that project to the hypothalamus are necessary for glucoprivic feeding and those that project spinally are necessary for glucoprivic control of adrenal medullary secretion. In the present study, we injected DSAP or control solution into the paraventricular nucleus of the hypothalamus (PVH) to investigate the role of NE/E neurons in glucoprivic control of glucocorticoid secretion. DSAP lesions significantly attenuated the magnitude and duration of the glucocorticoid (cortisol) response to 2-deoxy-D-glucose (2DG)-induced glucoprivation, but did not reduce the glucagon response. After 2DG (250 mg/kg), cortisol levels peaked at only 184% of pre-2DG levels in DSAP rats, compared to 440% in controls. Quantitative analysis revealed that DSAP did not destroy CRF-immunoreactive cell bodies in the PVH or terminals in the arcuate/median eminence, but did reduce dopamine β-hydroxylase immunoreactivity in hypothalamus and in hindbrain NE/E cell groups known to innervate the hypothalamus, including those that innervate CRF neurons in the PVH. Results indicate a critical role for hindbrain NE/E neurons in eliciting multiple controls of glucose homeostasis.
Related Products: Anti-DBH-SAP (Cat. #IT-03)
Bugarith K, Ritter S, Dinh T (2001) Hyperphagia and obesity results from the injection of the immunotoxin neuropeptide Y (NPY)-saporin (NPY-sap) into the paraventricular hypothalamus (PVH) of rats. Neuroscience 2001 Abstracts 948.2. Society for Neuroscience, San Diego, CA. Summary: NPY is a peptide implicated in the control of numerous physiological processes. A variety of G-protein coupled receptors, Y1, Y2, Y4, and Y5, mediate the actions of NPY. Recently a new immunotoxin, NPY-SAP, has been developed that is potentially of great importance for the study of NPY function. NPY-SAP is a conjugate of the peptide NPY and saporin, a plant toxin that inactivates ribosomes. The proposed mechanism of toxicity involves immunotoxin binding NPY receptors and destruction of neurons containing these receptors. In this study we injected different doses of NPY-sap or unconjugated saporin (SAP) into the PVH to test the effects of this new toxin. We found that at low doses, there was no effect of NPY-SAP on cytoarchitecture or immunoreactivity of select peptides in the PVH. There was also no difference in body weight between the groups. At high doses, there was histologically-detectable damage in the hypothalamus of NPY-SAP animals. We also found a major difference in body weight between the NPY-SAP rats and SAP controls. Ten weeks after the injections, the NPY-SAP rats (654.3g ± 39.04g) were much heavier than the SAP rats (410.6g ± 15.29g). Further, daytime (0800 - 1700H) food intake was about twice as much in NPY-SAP (9.53g ± 0.996g) as in SAP (5.74g ± 0.476g) rats, with no difference in overnight (1700 - 0800H) feeding. Work is ongoing to determine the specificity of the lesion and the mechanism of action of NPY-SAP. Related Products: NPY-SAP (Cat. #IT-28)
Bcl-2 improves survival of lesioned cholinergic neurons.
Lee KY, Leos R, Borowski TB, de Lacalle S (2001) Bcl-2 improves survival of lesioned cholinergic neurons. Neuroscience 2001 Abstracts 803.6. Society for Neuroscience, San Diego, CA.
Summary: Alzheimer’s disease is associated with the progressive cell death of cholinergic neurons in the central nervous system (CNS). A current challenge is to develop therapeutic approaches that could prevent atrophy and loss of CNS neurons, and promote regeneration of their processes. The purpose of our study is to determine the therapeutic potential of the antiapoptotic gene B-cell lymphoma 2 (bcl-2) on the cholinergic system of the basal forebrain (BF). Retrograde degeneration in the BF of 3-month-old rats was induced by unilateral injection of the immunotoxin 192 IgG-saporin into the entorhinal cortex. Fifteen minutes later, an injection of 5 µg of the pa22bgala4bcl-2 plasmid was placed into the BF. Similar procedures were followed for BF injections of a control plasmid or vehicle alone. We found that a single injection of bcl-2 plasmid into the BF of the lesioned side increased survival of cholinergic neurons ~50%, as compared to vehicle controls. In addition, cell survival was close to 100% of intact side with bcl-2 injections, compared to a 50% cell loss in animals injected with a control DNA. Our results indicate that an injection of the bcl-2 gene into the BF prevents loss of cholinergic neurons that have been injured by IgG-saporin. Mechanisms by which bcl-2 may protect cholinergic neurons could include retrograde transport of the plasmid to the nucleus and subsequent neuroprotective effects of increased levels of bcl-2 protein. Our results, in conjunction with other studies, suggest that neurodegeneration might be amenable to gene therapy.
Related Products: 192-IgG-SAP (Cat. #IT-01)
Bose P, Wang DC, Parmer R, Wiley RG, Thompson FJ (2001) Monoamine modulation of spinal reflex excitability of the lower limb in the rat: Intrathecal infusion (i.t.) of anti-DBH saporin toxin – time course for behavior. Neuroscience 2001 Abstracts 771.3. Society for Neuroscience, San Diego, CA.
Summary: Progressive neurophysiological changes in the excitability of ankle extensor stretch reflexes were observed following T8 spinal cord contusion injury. Our previous study indicated that nonspecific monoamine depletion (reserpine, i.p.)contributed to pathologic hyperreflexia. To test a more specific hypothesis, a longitudinal study was performed to evaluate the time course of changes in reflex excitability after i.t. injection of 250ng of anti-DBH saporin toxin (that specifically lesions descending spinal noradrenergic neurons) into the lumbar spinal cord of normal rats. Measures of ankle torque and time-locked EMGs were used to scale stretch reflex excitability across a broad range of stretch velocities (49-612°/sec) before and at weekly intervals following i.t. injection of toxin (n=12) or vehicle (n=6) using instrumentation and protocol previously reported. An elevated pattern of ankle toque was noted in all velocities tested on day 1 and week 1 of toxin treated animals compared with vehicle controls. By week-2, and for the remaining 5 weeks of testing, significant elevation of the ankle torque was only observed in the faster velocities. Significant increases in hindlimb axis and base of support were also observed from footprint analysis. These findings indicate that selective lesion of spinal noradrenergic fibers produced some of the specific changes in the reflex excitability that were observed following midthoracic spinal contusion injury.
Related Products: Anti-DBH-SAP (Cat. #IT-03)
Wang DC, Bose P, Parmer R, Wiley RG, Thompson FJ (2001) Monoamine modulation of spinal reflex excitability of the lower limb in the rat: Intrathecal (i.t.) infusion of anti-DBH saporin toxin – time course for neurophysiology. Neuroscience 2001 Abstracts 771.5. Society for Neuroscience, San Diego, CA.
Summary: Progressive neurophysiological changes in the excitability of ankle extensor stretch reflex were identified following T8 spinal cord contusion injury (Thompson et. al., 1992). Previous study suggested that nonspecific monoamine depletion may be a significant contributor to the pathologic hyperreflexia associated with chronic spinal cord injury (Thompson et. al., 1999). To test a more specific hypothesis, a study was performed to evaluate the time course of changes in reflex excitability after i.t. injection of 250ng of anti-DBH saporin toxin to specifically lesion descending spinal noradrenergic neurons into the lumbar spinal cord of normal rats. Measures of H-reflex excitability were obtained prior to and at weekly intervals following toxin injection until a physiological plateau was observed. Significant decreases in rate-depression of H-reflexes were observed by the second week after toxin infusion and were maintained throughout the five weeks of testing. These studies indicate that selective lesioning of noradrenergic fibers produced specific changes in reflex excitability previously observed following midthoracic spinal cord contusion injury in the rat. The results of this study further implicate neurophysiological changes associated with monoamine loss as a contributing factor leading to hyperreflexia derived from chronic spinal cord injury. Correlated behavioral changes are reported in the companion poster, Bose et.al.
Related Products: Anti-DBH-SAP (Cat. #IT-03)
Kohls MD, Majer KA, Russell BJ, Han Q, Blakely RD, Lappi DA (2001) A monoclonal antibody to an extracellular domain of the serotonin transporter: Characterization and targeting properties. Neuroscience 2001 Abstracts 814.9. Society for Neuroscience, San Diego, CA. PMID: 0
Summary: Using a peptide corresponding to a sequence from an extracellular domain of the rat serotonin re-uptake transporter (SERT), we have created a monoclonal antibody that recognizes the molecule on the cell surface. In FACS analysis, the antibody is able to recognize SERT-expressing human platelets and rat basophilic RBL-2H3 cells. Recognition is inhibited by the peptide immunogen. In western blotting of SERT-transfected cells, the antibody recognizes an approximately 95 kDa band corresponding to SERT, and shows no such recognition in the parent cell line. We have created an immunotoxin with this antibody by conjugation to the ribosome-inactivating protein saporin. This immunotoxin is 50-fold more cytotoxic than saporin to RBL-2H3 cells, according to ED50. The ED50 for the immunotoxin is 10.6 nM, whereas the ED50 for non-conjugated saporin is 477 nM. At 100 nM, the immunotoxin eliminates 72% of cells, whereas saporin alone has no significant difference on cells from control (addition of vehicle). These data demonstrate that antibody binding to SERT results in internalization, a situation we have also seen with the dopamine transporter (R.G. Wiley, M.B. Harrison, A.I. Levey and D.A. Lappi, submitted). Antibodies for the targeting and delivery of molecules to the interior of cells can be created through the use of peptide immunogens derived from the sequences of the extracellular domains.
Related Products: Antibody to Serotonin Transporter (SERT, Cat. #AB-N09)
Motooka Y, Kondoh T, Nomura T, Tamaki N, Nishizaki T (2001) Poster: Selective cholinergic denervation inhibits expression of long-term potentiation in the adult but not infant rat hippocampus. Neuroscience 2001 Abstracts 599.7. Society for Neuroscience, San Diego, CA.
Summary: The present study assessed the role of the cholinergic systems on the expression of perforant path long-term potentiation (LTP) in rat hippocampal slices from the infant and adult brain. To denervate the cholinergic systems, 192 1gG-saporin was injected into the lateral ventricle of the infant (2 weeks old) and adult (6 weeks old) rat brain. There, choline acetyltransferase immunoreactive fibers were little detectable two weeks and two months after injection for both the groups. For the infant rats, perforant path LTP was not affected by selective cholinergic denervation; the probability of LTP development was 0.83 (5/six slices) and 0.78 (7/nine slices) at 2 weeks and 2 months later in 192 IgG-saporin-treated slices, as compared with 0.83 at each period in control saline-treated slices. In contrast, the expression of the LTP was blocked by selective cholinergic denervation for the adult rats; the probability of LTP development was 0 (0/ten slices) and 0.38 (3/eight slices) at 2 weeks and 2 months later in 192 IgG-saporin-treated slices, as compared with 0.8 (8/ten slices) and 0.83 (5/six slices) at each period in control saline-treated slices. The results of the present study thus suggest that the cholinergic systems play a crucial role in the expression of LTP in the adult brain and that the denervated systems in the infant brain could be compensated by the sprouting of non-cholinergic fibers.
Related Products: 192-IgG-SAP (Cat. #IT-01)
Peterson WE, Jordan LM, Brownstone RM (2001) Immunolesioning of identified motoneuron pools following intramuscular injection of the immunotoxin, 192-IgG-saporin, in neonatal rats. Neuroscience 2001 Abstracts 626.14. Society for Neuroscience, San Diego, CA.
Summary: Studies have shown that the immunotoxin, 192-IgG-Saporin, can selectively lesion p75-positive cholinergic neurons of the basal forebrain in adult rats. Here we demonstrate the novel use of 192-IgG-Saporin to induce MN loss following intramuscular (I.M.) injection in neonatal rats. Two days following I.M. injection of 192-IgG-Cy3, neonatal rats (but not adult rats or neonatal mice) had Cy3-labeled MNs. This suggests that the 192-IgG antibody and its conjugates can be internalised by receptor-mediated endocytosis and retrogradely transported to spinal motor neurons. To induce MN loss, the left hind limb musculature of anaesthetised Sprague-Dawley rats were exposed, and several muscles injected with 0.5μg of 192-IgG-Saporin (Chemicon). Right hind-limb muscles were injected with DiI. Animals were sacrificed 25 days later. Ten μm coronal sections were obtained using a cryostat and Nissl stained. The neonatal rats showed signs of a locomotor deficit 2.5 weeks post injection with 192-IgG-Saporin, which increased slightly in severity over the next week and a half. Nissl stained coronal sections of the lumbar region showed an obvious MN deficit on the 192-IgG-Saporin treated side compared to control side. The injected muscles were also severely atrophic, a not unexpected finding given that they too express p75 receptors. We conclude that 192-IgG-Saporin can be used to lesion MN pools when IM injected in neonatal rats. This model may prove useful for testing cell replacement therapies for the treatment of MN diseases like amyotrophic lateral sclerosis (ALS).
Related Products: 192-IgG Mouse Monoclonal, Cy3-labeled (Cat. #AB-N43FL3)
Kudoh M, Seki K, Watanabe S, Shibuki K (2001) Sound sequence discrimination requires cholinergic inputs and suppression of M-currents in the rat auditory cortex. Neuroscience 2001 Abstracts 621.2. Society for Neuroscience, San Diego, CA.
Summary: Synaptic potentiation after sequential heterosynaptic stimulation is dependent on the stimulus sequence in the auditory cortex. Atropine or antagonists of M1 receptors block this sequence dependence. However, it is rescued by linopirdine, an inhibitor of M-currents, even in the presence of atropine. In the present study, we investigated the role of cholinergic inputs and the resulting suppression of M-currents in sound sequence discrimination. Rats were trained to discriminate sequence of two sounds. Licking a spout during sound presentation of a particular sequence was rewarded with water. The sounds of the rewarded or unrewarded sequence were randomly presented in a trial, which was repeated every one minute for 12 hours in 4 days. The percentage of the trials with a licking response to sounds was calculated separately for the rewarded and unrewarded sequence. Test performance, estimated from the difference, was significantly increased in the 4 days. Atropine (10 mg/Kg, i.p.) suppressed the increase of test performance. A cholinergic immunotoxin, 192IgG-saporin, was injected into the auditory cortex (62-400 ng/1µl) 1 week before the test. Sound sequence discrimination, but not discrimination between the two sounds, was significantly suppressed in these rats. Linopirdine (5 mg/Kg, i.p.) rescued the test performance of sound sequence discrimination in the rats injected with 192IgG-saporin. These results suggest that sound sequence discrimination requires cholinergic inputs and the resulting suppression of M-currents in the auditory cortex.
Related Products: 192-IgG-SAP (Cat. #IT-01)
Nattie EE, Li A (2001) Destruction by SP-SAP of rat retrotrapezoid nucleus (RTN) neurons expressing the neurokinin 1 receptor (NK1R) decreases breathing at rest and in response to hypercapnia. Neuroscience 2001 Abstracts 573.1. Society for Neuroscience, San Diego, CA.
Summary: Neurons in the RTN are hypothesized to provide both a tonic excitation for breathing and one location for central chemoreception (see Nattie, E., Prog. Neurobiol.1999). Lesions in anesthetized animals support the former while both lesions and focal acidification in unanesthetized animals support the latter. Application of substance P (SP) in the RTN increases respiratory output and immunohistochemistry for the SP (NK1) receptor shows extensive staining in the RTN. To destroy specifically these RTN neurons with NK1Rs we injected unilaterally in the RTN of the rat SP conjugated to the ribosomal toxin, saporin (SP-SAP; 100 nl; 1 uM; Advanced Targeting Systems). We measured ventilation by whole body plethysmography in the unanesthetized rat. At 6 to 15 days following SP-SAP injection, ventilation during air breathing was reduced by 19 to 24% and the response to 7% CO2 inhalation was reduced by 22 to 30%. Subsequent immunohistochemistry showed dramatically reduced NK1R staining in the area of the SP-SAP injection, which is difficult to quantify given the small number of RTN neurons and the extensive NK1R distribution along neuronal processes. RTN neurons with NK1Rs provide both a tonic excitation for breathing and a portion of the response to systemic hypercapnia.
Related Products: SP-SAP (Cat. #IT-07)
Sanders NM, Dinh TT, Pedrow C, Ritter S (2001) Selective immunotoxin lesions of hindbrain norepinephrine/epinephrine (NE/E) neurons impair feeding and corticosterone responses and Fos-immunoreactivity in hypothalamic sites during insulin-induced hypoglycemia (IIH). Neuroscience 2001 Abstracts 635.21. Society for Neuroscience, San Diego, CA.
Summary: Previously,we used the targeted immunotoxin saporin, conjugated to a monoclonal antibody against dopamine Beta-hydroxylase(DSAP),to destroy hypothalamic projecting NE/E neurons. Results showed that NE/E neurons are required for 2-deoxy-D-glucose induced feeding and Fos expression in the hypothalamus. In the present study,we used this same technique to determine if NE/E neurons play a similar role in mediating IIH responses.Rats were injected with DSAP or unconjugated saporin (SAP)into the hypothalamic paraventricular nucleus(PVH).Insulin reduced blood glucose to similar values in DSAP and SAP rats(15 and 17mg/dl, respectively). Glucagon responses to hypoglycemia were unaffected by DSAP,peaking at 597% and 504% of pre-drug levels after insulin in DSAP and SAP rats,respectively.In contrast,the corticosterone response was severely diminished in DSAP rats,peaking at only 123% of pre-insulin levels, compared to 353% in SAP rats.DSAP injections also abolished the feeding response to IIH.DSAP rats ate 0.9g of food during IIH while the SAP rats at 6.1g of food.DBH-ir was abolished in the A1/C1 overlap and reduced in A2,C2,C3 and A6 sites in DSAP rats. In the SAP rats,IIH induced Fos-ir in hindbrain NE/E neurons the PVH, LH and ARC.In DSAP rats, Fos-ir was reduced or abolished in these hypothalamic sites but was preserved in the adrenal medulla.These findings further support the role of hindbrain NE/E neurons in transmitting information from hindbrain glucoreceptive sites to hypothalamic circuits coordinating feeding and neuroendocrine responses to glucose deficit.
Related Products: Anti-DBH-SAP (Cat. #IT-03)
Dermorphin-saporin conjugate relieves inflammatory pain after peripheral application.
Palecek J, Paleckova V, Willis WD (2001) Dermorphin-saporin conjugate relieves inflammatory pain after peripheral application. Neuroscience 2001 Abstracts 508.10. Society for Neuroscience, San Diego, CA.
Summary: Opioid receptors have been shown to exist in specific population of DRG neurons signaling nociceptive information from peripheral tissues. In our study, we attempted to selectively destroy these neurons by using a peripheral application of the mu opioid agonist Dermorphin conjugated to ribosome inactivating toxin Saporin (DERM-SAP, Advanced Targeting Systems) in order to alleviate inflammatory pain. Intraarticular or intraplantar injection of carrageenan or CFA was used to induce inflammation in rats. The DERM-SAP conjugate was injected into the inflamed area 12-48h later. Responses of the animals to mechanical and thermal stimuli were tested before and after the inflammation and up to 21 days after the DERM-SAP application. The rats developed heat hyperalgesia in the affected paw 24 hours after the intraarticular CFA injection. In the saline injected group the hyperalgesia persisted for up to 19 days, but in the DERM-SAP injected group the signs of hyperalgesia were improving from day 7. Also mechanical allodynia tested with a VF filament (1.1g) was alleviated in the DERM-SAP group. In the carrageenan group, the DERM-SAP treatment decreased the heat hyperalgesia and prevented the development of hyperalgesia after repeated carrageenan application, 21days after the DERM-SAP treatment. Postmortem evaluation with a specific antibody showed presence of saporin in the DRG neurons. Our results show that peripheral application of DERM-SAP relieves inflammatory pain and suggest that peripheral application of neuropeptides conjugated to cell toxins or other substances such as antisense probes could be a useful tool for treating pain of peripheral origin. Supported by NIH grants NS09743 and NS11253.
Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)
Greco MA, Salin-Pascual R, Gerashchenko D, Blanco-Centurion C, Shiromani PJ (2001) Effects of 192-saporin lesion of the basal forebrain on sleep homeostasis and adenosine receptor (A1) mRNA levels. Neuroscience 2001 Abstracts 523.12. Society for Neuroscience, San Diego, CA.
Summary: Adenosine is hypothesized to be a mediator of sleep since adenosine levels in the basal forebrain increase after wakefulness and decrease following sleep. The inhibitory effect of adenosine on wake-active cholinergic neurons is thought to be mediated by the A-1 receptor subtype. We hypothesized that if adenosine inhibition of cholinergic neurons takes place via A-1A receptors on cholinergic neurons, the elimination of cholinergic cells should affect sleep homeostasis. To test this hypothesis, 192-saporin was used to selectively lesion basal forebrain cholinergic cells. 48h baseline sleep was recorded from male Sprague Dawley rats. Subsequently, the rats were kept awake for 12h and 24h recovery sleep was recorded. 192-saporin (4 ug) was then administered ICV. The rats were again continuously recorded for 3 weeks after the injection, a 12h prolonged waking period and during a 24h recovery sleep period. Brain sections processed for visualization of A1 mRNA and/or immunohistochemistry revealed that both ChAT- and parvalbumin-positive cells contained A-1A mRNA. 192-SAP eliminated ChAT immunoreactive cells in the basal forebrain. There were no differences in sleep-wakefulness up to 3 weeks after drug administration, a finding consistent with previous reports. In addition, there were no changes in recovery sleep following prolonged waking in lesioned rats. These results indicate that the cholinergic basal forebrain groups are not the primary mediators of wakefulness or of sleep homeostasis. We suggest that the effects of adenosine are mediated via binding to non-cholinergic neurons.
Related Products: 192-IgG-SAP (Cat. #IT-01)
Basal forebrain cholinergic system: Cortical activation, sleep/waking EEG and evoked potentials.
Shafi R, Berntson GG, Sarter M, Saurer T, Spino M (2001) Basal forebrain cholinergic system: Cortical activation, sleep/waking EEG and evoked potentials. Neuroscience 2001 Abstracts 533.16. Society for Neuroscience, San Diego, CA.
Summary: The role of the basal forebrain cholinergic system in cognitive functions such as arousal, attention and memory has been well documented. The purpose of the present study was to further elucidate the role of the basal forebrain in regulating cortical states and processes that may underlie these functions. Selective lesions of the cholinergic neurons of the basal forebrain were made using the immunotoxin 192 IgG-saporin, which selectively targets the p75 receptor on cholinergic neurons. The effects of these lesions on sleep structure and EEG activity and on afferent priming of cortical reactivity was evaluated. Specifically, we monitored behavioral activity and sleep states and examined the frequency distribution of power distribution in EEG frequency bands during these states. In separate sessions, we also recorded cerebral event-related potentials to auditory stimuli (100 ms, 1 K Hz at 60, 70 and 80 db) after intraperitoneal administration of saline or epinephrine (0.5 mg/kg, which we have previously found to result in priming or enhancement of the auditory evoked response). Compared to controls, lesioned animals showed a reduction in spontaneous activity, reduced power in higher frequency (primarily gamma) EEG bands during both sleep and waking, and altered sleep structure. In addition, lesioned animals displayed lower amplitude auditory evoked potentials and a loss of epinephrine-priming of the evoked response. Results support the view that the basal forebrain cholinergic system may play an important role in cortical activation and the regulation of sleep/waking states, as well as in cortical processing and its enhancement by visceral priming.
Related Products: 192-IgG-SAP (Cat. #IT-01)
Zambon NJ, Nagle R, Pokala V, Gibbs RB, Johnson DA (2001) Low dose 192 IgG-saporin selectively destroys basal forebrain cholinergic neurons and impairs acquisition of a spatial memory task. Neuroscience 2001 Abstracts 534.13. Society for Neuroscience, San Diego, CA.
Summary: We previously showed that a high dose (1 μg) of the selective cholinergic immunotoxin 192 IgG-saporin (SAP), injected into the medial septum (MS) of Sprague-Dawley rats, impeded acquisition of a delayed matching-to-position (DMP) spatial memory task, whereas injections of ibotenate (5 μg in 1 μL) did not. The present study examined the effects of lower doses of SAP (0.22 and 0.45 μg in 1 μl) on DMP acquisition. Animals received either SAP or vehicle injected directly into the MS. Two weeks later, animals were food deprived and trained to the DMP task. Rats received 8 trial pairs/day until they reached a criterion of 15/16 correct choices. Seven days later, post-criteria testing for retention was performed. Brain tissues were analyzed for choline acetyltransferase (ChAT) activity, or were processed for immunohistochemical detection of ChAT and parvalbumin. Control rats required significantly fewer days (13.1) to reach criterion than rats that received 0.22 (22.0 days) or 0.45 (20.1 days) μg SAP. There was no effect of SAP treatment on post-criteria testing. Injections of SAP produced marked depletion of ChAT-positive cells and ChAT activity, but no apparent depletion of parvalbumin staining in the MS. In contrast, ibotenate injections used in the previous study were shown to produce marked depletion of parvalbumin staining in the MS, but no significant cognitive impairment. The data suggest that selective destruction of cholinergic neurons in the MS significantly impairs acquisition of the DMP task.
Related Products: 192-IgG-SAP (Cat. #IT-01)
Loss of histaminergic neurons does not produce hypersomnolence.
Chou TC, Gerashchenko D, Saper CB, Shiromani PJ (2001) Loss of histaminergic neurons does not produce hypersomnolence. Neuroscience 2001 Abstracts 522.21. Society for Neuroscience, San Diego, CA.
Summary: Electrolytic lesions of the posterior hypothalamus (PH) produce long-lasting hypersomnolence (1,2). The PH contains histaminergic neurons in the tuberomammillary nucleus (TMN) that project diffusely throughout the brain. Because histamine promotes wakefulness while antihistamines are sedating, the TMN is thought to be critically involved in maintaining wakefulness. To test this hypothesis, we placed cell-specific lesions in the PH and TMN of rats and measured sleep-wake behavior. Lesions were produced using either the conventional excitotoxin ibotenic acid, or the novel toxin orexin (hypocretin) conjugated to the ribosomal toxin saporin (ORX/HCRT-SAP). Ibotenic acid injections were ineffective at lesioning the TMN; most histaminergic neurons were selectively spared while neurons in surrounding regions such as the mammillary bodies and supramammillary area were completely lesioned. In contrast, ORX/HCRT-SAP injections into the TMN lesioned up to 95% of histaminergic neurons, as determined by adenosine-deaminase immunostaining, with a similar loss of neurons in adjacent areas. Surprisingly, neither group of rats showed changes in NREM or REM sleep time or circadian distribution of sleep relative to saline-injected controls for up to 2 weeks after surgery. Thus, the waking state may not be critically dependent on the PH or TMN in rats. Further research is needed to reconcile the sedating effects of antihistamines with the current findings. 1. Ranson 1939, Archiv Neurol and Psychiatry 41(1):1-23. 2. Swett and Hobson 1968, Arch Ital Biol 106(3):283-293.
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Sherren N, Pappas BA (2001) Behavioural and neurochemical changes associated with single and combined acetylcholine and dopamine lesions in neonatal rats. Neuroscience 2001 Abstracts 539.5. Society for Neuroscience, San Diego, CA.
Summary: The functional outcomes of neonatal ACh or DA lesions are frequently less severe or qualitatively different from those seen in adult rats, and may be due to compensatory neurochemical changes. Given that these transmitter systems interact in the adult brain and that ACh and DA hypofunction may underlie the cognitive and motor disabilities seen in Rett syndrome, we hypothesized that combined neonatal ACh/DA lesions may produce a profile of neurochemical changes and behavioural impairments which are more severe or distinct from that caused by either lesion alone. Rats were lesioned at postnatal day 7 with 192 IgG-saporin (ACh rats), 6-OHDA with NE receptor blockade (DA rats), or both (ACh/DA rats). Behavioural testing occurred at 4 months of age. In the open field, only ACh/DA rats exhibited locomotor hyperactivity whereas all lesioned groups exhibited reduced exploratory behaviour. Neither DA nor ACh/DA rats were able to solve the Morris water maze, however ACh rats were indistinguishable from controls. 192 IgG-saporin treatment produced a 75% decrease in hippocampal ChAT activity, and cortical decreases of 30%, 70% and 40% in the frontal/cingulate (FC), retrosplenial (RS) and partietotemporal (PT) regions respectively. 6-OHDA treatment produced a 90% decrease in striatal DA levels and a 75% decrease in FC cortex. Interestingly PT DA levels were 68% higher in ACh rats but 47% lower in ACh/DA rats compared to control, while DA rats showed a decrease which was not significantly different from control. Thus sparing of spatial learning ability in ACh rats may be mediated by increases in PT DA levels, whereas combined ACh/DA lesions exacerbate DA loss in this region.
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Immunolesioning of brainstem DBH neurons on the mating-induced LH and prolactin surge in the rabbit.
Pau K (2001) Immunolesioning of brainstem DBH neurons on the mating-induced LH and prolactin surge in the rabbit. Neuroscience 2001 Abstracts 466.7. Society for Neuroscience, San Diego, CA.
Summary: Coitus induces a surge release of norepinephrine (NE) that is accompanied by a preovulatory gonadotropin-releasing hormone (GnRH)/luteinizing hormone (LH) surge. Prazosin, an alpha-1 adrenergic antagonist, attenuates the GnRH/LH surge, and tyrosine hydroxylase (TH) gene expression in brainstem NE areas increases within 30 min after coitus. Here, we determined the coitus-induced LH/prolactin surge after specific lesioning of dopamine beta-hydroxylase (DBH) neurons in the brainstem with monoclonal anti-DBH sera conjugated with the ribosomal cytotoxin saporin (DBH-SAP). Female NZW rabbits received 3rd cerebroventricular injection (Day 0) of either DBH-SAP (20 µg, n=4) or SAP (3 µg, n=4). On day 14, the four DBH-SAP females were paired with stud males, but none of them mated. After daily injection of estradiol benzoate (EB, 3 µg) for 3 days, all eight females mated. Blood samples were taken once before, and at 10-min intervals for 4 hours after, coitus. Brainstems were prepared for immunocytochemical detection of DBH and TH. Coitus increased both LH and prolactin release in either DBH-SAP or SAP animals. However, postcoital LH and prolactin levels were 55% lower and 50% higher, respectively, in DBH-SAP rabbits than in SAP animals. The number of DBH neurons was near zero in the A6 and reduced by 80% in the A1 and 70% in the A2 noradrenergic areas in DBH-SAP animals. The number of TH neurons was reduced by 95% and 30% in the A6 and A1 areas, respectively, and did not change in the A2 area. The results suggest that the presence of intact brainstem NE neurons are critical for sexual performance and production of normal LH/prolactin surge after coitus in female rabbits.
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Neonatal cholinergic lesions alter reactivity to a GABAergic agonist in 18-day-old rats.
Ricceri L, Scattoni ML, Calamandrei G (2001) Neonatal cholinergic lesions alter reactivity to a GABAergic agonist in 18-day-old rats. Neuroscience 2001 Abstracts 541.14. Society for Neuroscience, San Diego, CA.
Summary: We have shown previously that neonatal intracerebroventricular (icv) injections of the selective cholinergic immunotoxin 192 IgG-saporin on postnatal day (pnd) 7induces learning impairments on pnd 15 and disruption of reactivity to spatial novelty on pnd 54. The same neonatal treatment also induces a permanent cholinergic loss in both hippocampus and neocortex. In the present study we analyzed behavioral effects induced by a GABAergic drug (muscimol, a GABAa receptor agonist) in rats neonatally lesioned with 192 IgG-saporin (icv on pnd 7). On pnd 18 192 IgG-saporin lesioned and sham rats were injected with muscimol (0.1, 0.5 mg/kg ip) and placed in an open field arena for 20 min; locomotion, wall rearing and rearing responses were measured. In sham animals, as expected, 0.1 muscimol decreased locomotion, wall rearing and rearing responses. In saporin lesioned animals 0.1 muscimol increased locomotion, left wall rearing responses unchanged and decreased only rearing responses. In a 60s hot-plate test, 0.1 muscimol induced comparable analgesic responses in both sham and saporin-lesioned animals. The 0.5 muscimol dose resulted cataleptic for both saporin and sham lesioned rats. Neonatal saporin per se also reduced wall rearing and rearing responses. These data suggest that only in selective behavioral patterns -- associated with locomotion and exploration of the environment -- reactivity to a GABAergic agonist is reduced following neonatal cholinergic lesions, probably because of a decrease of GABAa receptors in the medial septal nucleus.
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α-1 adrenergic agonist effect on cholinergic muscarinic receptors.
Harrell LE, Kolasa K, Parsons DS, Conger K (2001) α-1 adrenergic agonist effect on cholinergic muscarinic receptors. Neuroscience 2001 Abstracts 549.16. Society for Neuroscience, San Diego, CA.
Summary: Degeneration of the basal forebrain cholinergic system and sympathetic ingrowth appear to be pathological changes in Alzheimer's Disease patients (AD), leading to an alteration in the balance between both systems and may mediate cognitive deficits in AD. In an attempt to model this situation, intraventricular injection (ivc) of a specific cholinergic immunotoxin, 192-IgG-saporin, has been used to induce peripheral noradrenergic fibers to grow into cortex and hippocampus after cholinergic denervation of rat cortex (CCD) and hippocampus (HCD). This adrenergic reorganization has been termed cortical (CSI) and hippocampal (HSI) sympathetic ingrowth. 192-IgG-saporin ivc injection was followed by intraperitoneal (ip) treatment with α1 agonist methoxamine. Thus the effects on choline acetyltransferase (ChAT) activity, norepinephrine (NE) level and muscarinic acetylcholine receptors (mAChR) were studied in rat hippocampal and cortical brain tissue. We found that 192-IgG-saporin produced significant decrease in ChAT activity in all experimental groups and areas. Methoxamine (3 and 6 mg/kg ip) did not affect NE levels. It produced significant decrease in mAChR affinity in the cholinergic denervation group and no significant increase in mAChR density in cholinergically denervation groups of dorsal hippocampal and cortical areass. Results of the present study indicate the influence of α1 agonist treatment on mAChR and may provide new concepts for the future combination drug therapy for AD patients.
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α-1 adrenergic antagonist effect on cholinergic muscarinic receptors.
Kolasa K, Harrell LE, Parsons DS, Conger K (2001) α-1 adrenergic antagonist effect on cholinergic muscarinic receptors. Neuroscience 2001 Abstracts 549.17. Society for Neuroscience, San Diego, CA.
Summary: Cholinergic denervation of cortex and hippocampus in rat causes an unusual neuronal rearrangement, in which peripheral sympathetic fibers, originating from superior cervical ganglia, grow into the cholinergically denervated areas. This process has been termed cortical (CSI) and hippocampal sympathetic ingrowth (HSI). A similar process may occur in Alzheimer's Disease (AD). Recent studies suggest that the balance between central adrenergic and cholinergic systems may be important for normal learning and memory, while the alterations of these systems may play a critical role in cognitive deficits of AD. To better understand this situation specific cholinotoxin, 192-IgG-saporin, was intraventricularly (ivc) injected to produce a selective loss of cholinergic cells in rat basal forebrain nuclei, cholinergically denervating hippocampal (HCD) and cortical areas (CCD). This effect was confirmed by significant decrease in choline acetyltransferase activity in all groups and brain structures.192-IgG-saporin injection was followed by a treatment with α1-adrenergic antagonist prazosin to determine the effect on hippocampal and cortical muscarinic acetylcholine receptors (mAChR) and norepinephrine (NE) level. Prazosin (0.5 & 2 mg/kg ip) produced decreases in NE levels of HSI and CSI and induced no significant increase in mAChR affinity in HSI and CSI groups in dorsal hippocampus,anterior and entorhinal cortex. Injected at the dose of 2 mg/kg it increased mAChR density in CSI of both cortical areas. The present results began to define the interaction between adrenergic and cholinergic systems, as α1 antagonist treatment affects mAChR,a potential therapeutic target in AD.
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Wang H, Guyenet PG (2001) DL-homocysteic (DLH)-induced tachypnea is eliminated by ablation of neurokinin-1 receptor immunoreactive (NK1R-ir) neurons of the preBötzinger complex (preBötc). Neuroscience 2001 Abstracts 633.4. Society for Neuroscience, San Diego, CA.
Summary: We identified a hot spot of the ventrolateral medulla (VLM) where tachypnea is produced by small injections of the excitatory amino acid DLH in urethane-anesthetized vagotomized rats. We sought to determine its anatomical location relative to a group of NK1R-ir cells that may be a marker of the preBötC. We also determined the location of the hot spot relative to VLM pressor and depressor sites. The VRG was located by recording respiratory units and then 5-10 nl of 10 mM DLH were injected into the VRG at 200 mm interval from Bregma -12.1 to -13.3 mm. DLH produced site-specific changes in respiratory rate and mean arterial blood pressure (MAP). At rostral levels (Bregma -12.1 mm) PND rate decreased and MAP increased (-57 ± 11% and 9.4 ± 0.9%; N= 4-6). At more caudal levels these effects gradually reversed (cross-over point at -12.4 mm). Tachypneic responses were restricted between Bregma -12.5 and -13.1 mm with a sharp peak (88.7 ± 12.8% increase in rate) at -12.7 mm. The hot spot corresponded to where most pre-inspiratory neurons are found. It overlapped with VRG NK1R-ir cells but was more restricted than the distribution of these cells. Depressor responses were maximal at -12.7 mm and stable at more caudal levels. After unilateral ablation of the NK1R-ir cells (N=3) with a saporin-NK1R agonist conjugate, the DLH-induced tachypnea disappeared bilaterally. In conclusion, the tachypnea hot spot corresponds to the defined preBötC. It overlaps with the rostral end of the VLM depressor area and the NK1R-ir neurons of the preBötC may be responsible for DLH-induced tachypnea.
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Sarter M, Burk JA, Graf A, Russell J, Bruno JP (2001) Basal forebrain cholinergic and gabaergic neurons mediate different aspects of sustained attention performance in rats. Neuroscience 2001 Abstracts 313.7. Society for Neuroscience, San Diego, CA.
Summary: Several studies have demonstrated that lesions of basal forebrain (BF) corticopetal cholinergic neurons, produced by infusing the cholinotoxin 192IgG-saporin, impair the ability to detect visual signals in an operant sustained attention task but not to reject nonsignals. Information about the functions of BF non-cholinergic, particularly GABAergic neurons has remained scarce. As infusions of the excitotoxic amino acid ibotenic acid (IBO) are known to predominantly destroy non-cholinergic neurons in the BF, the effects of BF IBO lesions on sustained attention performance were assessed. Rats were trained to perform a sustained attention task and then received bilateral infusions of IBO (0.06 M; 0.5 μL / hemisphere) or of saline (N=8 /group) into the basal forebrain. Postsurgically, rats were trained to stable performance on the sustained attention task. Compared to controls, ibotenic acid-lesioned rats more frequently responded to non-signal events by 'claiming' a hit, and they exhibited an increase in hits in response to the briefest signals. Immunohistological analyses confirmed the predominant loss of parvalbumin-positive and thus presumably GABAergic neurons in the BF, while ChAT-positive neurons were partially spared. Likewise, IBO-lesioned animals exhibited an only moderate decrease in cortical AChE-positive fiber density. These and additional findings indicate that BF GABAergic neurons contribute to attentional performance by mediating the animals' ability to switch between the response-rules for signals and non-signals.
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Janczewski WA, Gray PA, Feldman JL (2001) Central origin of ataxic breathing after lesion of preBötzinger complex (preBötc) neurokinin 1 receptor expressing (NK1R+) neurons. Neuroscience 2001 Abstracts 243.2. Society for Neuroscience, San Diego, CA.
Summary: Pathological breathing results from near complete lesions of preBötC NK1R+ neurons in awake adult rats (Gray et al., FASEB J.,15, 2001). To determine whether this ataxic pattern is central in origin, we examined the breathing pattern of rats using combined diaphragmatic EMG (diaEMG) and whole body plethysmography (WBP). Under anesthesia, substance P conjugated to saporin was injected bilaterally into the preBötC (n=7) and EMG electrodes were implanted into the diaphragm (n=10). Up to four days postinjection, all rats breathed normally. DiaEMG postinpiratory activity was evident in all rats and accentuated during brief apnea following spontaneous sighs. After postinjection day 5, injected rats showed a transformation in breathing pattern from normal to ataxic characterized by high frequency breaths of varying amplitude separated by periods of tonic diaphragmatic discharge. There was no lag between the WBP output and diaphragmatic activity (WBP measures virtually paralleled the moving average of diaEMG activity), suggesting the absence of significant flow limitations. During apnea, nonrespiratory movement produced artifacts on WBP signal but did not affect diaEMG. We conclude that ablation of preBötC NK1R+ neurons leads to hypoventilation and apneas of purely central origin without upper airway obstruction or bronchoconstriction.
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Cholinergic and noncholinergic septal neurons modulate strategy selection in place learning.
Cahill JF, Baxter MG (2001) Cholinergic and noncholinergic septal neurons modulate strategy selection in place learning. Neuroscience 2001 Abstracts 314.14. Society for Neuroscience, San Diego, CA.
Summary: Rats solving a simple spatial discrimination task in a plus-maze initially employ a place learning strategy, then switch to a motor response strategy. The hippocampus is required for the use of a place learning strategy in this task. Rats with 192 IgG-saporin lesions of the medial septum/vertical limb of the diagonal band (MS/VDB), that selectively removed cholinergic neurons projecting to the hippocampus, were significantly facilitated in acquisition of the spatial discrimination, and switched from place to response strategies just as control rats did. Rats with ibotenic acid lesions of the MS/VDB, which produced cell loss in the MS/VDB but little damage to cholinergic neurons, were significantly impaired in acquiring the spatial discrimination and did not reliably employ either a place or response strategy at any point in training. This suggests that the MS/VDB modulates hippocampal involvement in place learning, but that cholinergic MS/VDB neurons are neither necessary nor sufficient for using a place strategy to solve a spatial discrimination.
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Environment-spatial conditional learning: Contribution of medial septal cholinergic neurons.
Jackson O, Firoz EF, Janisiewicz AM, Baxter MG (2001) Environment-spatial conditional learning: Contribution of medial septal cholinergic neurons. Neuroscience 2001 Abstracts 314.15. Society for Neuroscience, San Diego, CA.
Summary: Visual-spatial conditional discrimination learning is impaired by damage to the cholinergic septohippocampal neurons in marmoset monkeys (Ridley et al., 1999). We sought to explore the generality of this finding by testing rats with selective lesions of cholinergic septohippocampal projections (made with 192 IgG-saporin) on an environment-spatial conditional discrimination task. In this task, one of two sets of local environmental cues (consisting of a unique geometric shape with unique visual stimuli) directed search to a particular goal location in the environment (selected from eight possible locations). Preliminary observations suggest that rats with selective lesions of medial septal cholinergic neurons are impaired on acquiring this conditional discrimination task, but are unimpaired on acquiring a single discrimination problem using the same cues. This finding is consistent with a general role for septohippocampal cholinergic projections in the learning of conditional discrimination problems, suggesting that medial septal cholinergic neurons subserve cognitive functions other than decremental attentional processing (Baxter et al., 1997, 1999).
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Effects of basal forebrain cholinergic lesions on spatial learning in male and female rats.
Rodefer JS, Jonasson Z, Cahill JF, Tobey RE, Baxter MG (2001) Effects of basal forebrain cholinergic lesions on spatial learning in male and female rats. Neuroscience 2001 Abstracts 314.16. Society for Neuroscience, San Diego, CA.
Summary: Studies with a selective toxin for basal forebrain cholinergic neurons have consistently found little spatial learning impairment in rats with lesions limited to basal forebrain cholinergic neurons. However, the basal forebrain cholinergic system is extensively modulated by estrogen and related sex hormones. Furthermore, female mice are more susceptible than male mice to scopolamine-induced impairments in water maze performance (Berger-Sweeney et al., 1995). Hence, behavioral effects of selective cholinergic lesions may differ between male and female rats, a possibility that has not been directly assessed experimentally. In the present study, male and female Long-Evans rats were given injections of 192 IgG-saporin into the medial septum/vertical limb of the diagonal band (MS/VDB), or a control surgery, and postoperatively tested on several spatial learning tasks in the Morris water maze. On place acquisition, female control rats performed worse than male controls; MS/VDB-lesioned rats performed identically to male controls, regardless of sex. Male rats performed better than female rats on reversal of place discrimination, regardless of lesion status. In a test of place learning strategy, cholinergic lesions appeared to enhance the use of a spatial (vs. a motor response strategy) in rats of both sexes. Hence, female rats are not more susceptible to place learning impairment following loss of cholinergic input to the hippocampus; indeed these lesions seem to enhance place learning in female rats. Furthermore, removal of cholinergic input to the hippocampus seems to enhance the selection of a spatial strategy.
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Bartness TJ, Demas GE (2001) A novel method for localized sympathetic nervous system denervation of peripheral tissue using guanethidine. Neuroscience 2001 Abstracts 309.10. Society for Neuroscience, San Diego, CA.
Summary: A simple technique for the functional deactivation of the sympathetic nervous system innervation of peripheral tissues is described using the local application of guanethidine. Multiple unilateral microinjections of guanethidine were made into one inguinal or epididymal white adipose tissue (IWAT and EWAT) pads of hamsters, whereas the contralateral pad received equivolumetric saline vehicle injections. Guanethidine treatment virtually abolished the sympathetic innervation of both EWAT and IWAT, as measured by the absence of significant norepinephrine (NE) tissue content two weeks later and as suggested by the two-fold increase in IWAT mass characteristic of surgically induced WAT denervation. IWAT and EWAT NE content and mass were unaffected in the contralateral control pads. Guanethidine injections into the spleen also lead to a function sympathectomy as indicated by significant depletions of NE content. Because guanethidine treatment did not decrease body mass nor induce ptosis, no chemical-induced malaise or global sympathetic denervation, respectively, was suggested. We compared the effects of local guanethidine treatment on IWAT NE content and pad mass with the local application of the sympathetic neurotoxin, anti-dopamine beta hydroxylase saporin, and with local surgical IWAT denervation. Guanethidine treatment significantly reduced IWAT NE content to a greater degree than for the alternative sympathectomy methods. These results suggest that locally applied, chemical sympathectomy with guanethidine provides an effective, restricted method for denervating WAT and likely other peripheral tissues.
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Butt AE, Noble MM, Barrett H, Brinegar S, Hixon A, Kester B, Levey D, Rea T, Ryan M, Schulze S, Vogel L (2001) 192 IgG saporin lesions of the nucleus basalis magnocellularis impair working but not reference memory in a delayed non-match-to-position y-maze task. Neuroscience 2001 Abstracts 314.17. Society for Neuroscience, San Diego, CA.
Summary: The current experiment tests the hypothesis that the cortically projecting cholinergic neurons of the nucleus basalis magnocellularis (NBM) are selectively involved in working and not reference memory. Male Long-Evans rats received extensive pre-operative training in a delayed non-match-to-position (DNMTP) task in a Y-maze. In this task, 0-s, 1-min, or 2-min delays were interposed between consistently food-reinforced “sample runs”, where rats were allowed access to only one arm of the maze, and “choice runs”, where rats were allowed access to both arms of the maze but were reinforced only for entering the arm opposite that entered on the previous sample run. Upon completion of pre-operative training, rats were matched for performance and were randomly assigned to a sham lesion group or to the NBM lesion group, which received bilateral infusions of the cholinergic immunotoxin 192 IgG-saporin into the NBM. As predicted, post-operative performance in the NBM lesion group was normal at the 0-s delay but was severely impaired at the 1-min and 2-min delays. Results suggest that the NBM is critically involved in supporting normal spatial working memory but that previously acquired reference memory for the non-matching rule remains intact following selective NBM lesions.
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Estacio MA, Tsukamura H, Reyes BA, Maeda KI (2001) Noradrenergic inputs to the paraventricular nucleus (PVN) is involved in estrogen receptor α expression in the PVN of 48-h fasted and 2DG-injected female rats. Neuroscience 2001 Abstracts 409.6. Society for Neuroscience, San Diego, CA.
Summary: Involvement of noradrenergic inputs to the paraventricular nucleus (PVN) in estrogen receptor α (ERα) expression in the PVN during 48-h fasting and 2DG-induced glucoprivation in female rats was determined by examining the effect of destroying the noradrenergic inputs to the PVN using the saporin-conjugated anti-dopamine-β-hydroxylase (anti-DBH-saporin). Ovariectomized rats were injected bilaterally with anti-DBH-saporin in the PVN. After two weeks, animals were either fasted for 48 hours or injected intravenously with 2DG, then perfused with 4% paraformaldehyde. Brain sections were processed for ERα and DBH immunocytochemistry. Forty-eight-hour fasting or 2DG injection siginificantly increased the number of ERα-immunoreactive (ERα-ir) cells in the PVN in control animals. Anti-DBH-saporin injection prevented fasting- or 2DG-induced increase in ERα-ir cells in the PVN. The DBH-ir axons in the parvocellular PVN were severely reduced following anti-DBH-saporin injection in both fasted and 2DG injected rats. Among the brainstem noradrenergic cell groups examined, there was a significant decrease in the number of DBH-ir cells in the A2 region of both fasted and 2DG injected rats treated with anti-DBH-saporin. There was no obvious reduction in the number of DBH-ir cells in the A1 and A6 regions in the anti-DBH-saporin-injected fasted- or 2DG-injected rats. The results suggest that the A2 noradrenergic input to the PVN plays a major role in increasing ERα expression in the PVN in response to 48-h fasting or 2DG-induced glucoprivation.
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Dose-dependent effects of intrathecal substance P-saporin and SSP-saporin.
Wiley RG, Kline IV RH, Lappi DA (2001) Dose-dependent effects of intrathecal substance P-saporin and SSP-saporin. Neuroscience 2001 Abstracts 281.11. Society for Neuroscience, San Diego, CA.
Summary: Selective destruction of lamina I dorsal horn neurons expressing the neurokinin-1 receptor (NK-1R) can attenuate responses to capsaicin injection and thermal hyperalgesia/mechanical allodynia in models of inflammatory, persistent or neuropathic pain. In the present study, we sought to determine the relationships between spinal intrathecal dose of substance P-saporin or the related toxin, SSP-saporin, the loss of NK-1R neurons and reduction of phase II formalin responses. Rats were injected intrathecally with 10 ul of either vehicle, 175 ng, 350 ng or 700 ng of SP-sap. Others were injected with either vehicle, 25 ng, 50 ng or 100 ng of SSP-sap. After 2 weeks, nocifensive behavior was scored for 90 min after a unilateral hindpaw injection of dilute formaldehyde. The amount of phase II nocifensive behavior from 20-90 min post injection was totaled for each animal. Rats were sacrificed and transverse lumbosacral spinal cord sections were stained for NK-1R using indirect immunoperoxidase technique. Digital micrographs of the superficial dorsal horn were captured and the number of pixels in the darkest intensity values were expressed as percent of the analysis area for each dorsal horn. Significant correlations were noted for dose vs dark pixel percentage and for dark pixel percentage vs phase II formalin behavior. The greater the toxin dose the greater the loss of NK-1R staining and the greater the attenuation of phase II formalin behavior. These results indicate that the toxin effects on pain behavior are proportional to the degree of loss of lamina I NK-1R expressing neurons.
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The role of the cholinergic basal forebrain in learning, memory and reward expectancies.
Savage LM, Buzzette R, Ramirez D (2001) The role of the cholinergic basal forebrain in learning, memory and reward expectancies. Neuroscience 2001 Abstracts 314.18. Society for Neuroscience, San Diego, CA.
Summary: The cholinergic basal forebrain degenerates in Alzheimer's Disease and the degree of this degeneration correlates with a decline in cognitive processing. In the present study we have modeled this degeneration in the rat by the selective immunotoxin 192 IgG-Saporin. This immunotoxin destroys cholinergic neurons in the basal forebrain nuclei in rats and thus allows for the study of the impact of cholinergic deafferentation on learning, memory, and other cognitive processes without direct effects on other neuronal systems. After intracerebroventricularly infusions of the immunotoxin or vehicle solution, male rats were allowed to recover for three weeks before being tested in a matching-to-position task. The matching-to-position task was altered to influence the type of cognitive strategies a subject would use to solve the task. The main behavioral manipulation was the use of the differential outcome procedure (DOP). The DOP involves correlating each to-be-remembered event with a distinct reward condition. We found that cholinergic lesions did not dramatically impair learning the matching rule. However, the memory performance of subjects with cholinergic lesions was dramatically impaired – if subjects were not trained with the DOP. When subjects were trained with the DOP, and relied on reward expectancies to solve the delayed-matching-to-position task the cholinergic lesion had little effect. These findings demonstrate that cholinergic immunolesions by 192IgG-saporin induce specific cognitive impairments—dependent on task demand characteristics.
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Hamaue N, Yamazaki Y, Ohmori H, Sumikawa K (2001) Nicotine enhances N-methyl-D-aspartate receptor responses and facilitates long-term potentiation in the hippocampus from rats with cholinergic lesions. Neuroscience 2001 Abstracts 376.5. Society for Neuroscience, San Diego, CA.
Summary: Nicotine reverses cognitive impairments caused by lesion of the cholinergic system and improves performance of Alzheimer's patients. The mechanisms underlying these effects of nicotine, however, are unknown. Because nicotine facilitates the induction of N-methyl-D-aspartate receptor (NMDAR)-dependent long-term potentiation (LTP) in the hippocampal CA1 region, we examined whether nicotine enhances NMDAR responses and facilitates LTP induction in the hippocampus from rats with cholinergic lesions. Selective cholinergic denervation of rat hippocampus was performed by the immunotoxin 192 IgG-saporin. We then recorded burst NMDAR responses in hippocampal slices prepared from 192-IgG-saporin-treated rats and found that nicotine (1 μM) enhanced burst NMDAR responses. When GABAergic transmission was completely blocked by picrotoxin, nicotine had no effect on burst NMDAR responses. We also monitored the induction of LTP in 192-IgG-saporin-treated hippocampi and found that a weak tetanus (20 pulses at 100 Hz), which induced LTP in PBS-treated hippocampi, failed to induced LTP. However, in the presence of nicotine (1 μM), a same weak tetanus induced LTP in 192-IgG-saporin-treated hippocampi. Our results suggest that nicotine potentiates NMDAR responses by disinhibition of pyramidal cells and facilitates LTP induction in the hippocampus from animals with cholinergic lesions. The observed nicotine effects may represent the cellular mechanism underlying the compensatory action of nicotine in the presence of cholinergic deficits.
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Maze strategy in rats with GABAergic or cholinergic lesions of medial septum.
Yoder RM, Reuss SA, Pang KC (2001) Maze strategy in rats with GABAergic or cholinergic lesions of medial septum. Neuroscience 2001 Abstracts 418.7. Society for Neuroscience, San Diego, CA.
Summary: Cholinergic and GABAergic neurons are the two major cell types that project from medial septum to hippocampus. Although complete lesions of hippocampus or medial septum impair spatial memory, selective lesions of cholinergic or GABAergic neurons do not impair spatial abilities on an 8-arm radial or Morris water maze. Control and lesion rats possibly use different strategies to solve these tasks. Previous studies show that normal rats initially use a place strategy, then switch to a response strategy. In contrast, rats with hippocampus inactivated rely on a response strategy. Our preliminary results suggest that rats with GABAergic lesions of medial septum preferentially use a response strategy. The present study assessed whether rats with cholinergic (192-IgG saporin) or GABAergic (kainic acid) lesions of the medial septum preferentially use a place, response, or cue strategy to solve a plus maze task. During training, one arm contained food (goal) and an adjacent arm served as the starting location. The room contained distal cues and a proximal cue near the food. Probe trials were used every 4th day to assess the maze strategy used by the rat. During probe trials, the start location was located on the arm opposite the original start arm, and the proximal cue was located in the original start location. This study determines whether rats with loss of cholinergic or GABAergic medial septal neurons preferentially use different strategies to solve maze tasks.
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Selective neonatal neurochemical lesions persist into old age and cause Alzheimer-like pathology.
de Butte M, Fortin T, Sherren N, Pappas BA (2001) Selective neonatal neurochemical lesions persist into old age and cause Alzheimer-like pathology. Neuroscience 2001 Abstracts 426.8. Society for Neuroscience, San Diego, CA.
Summary: There is a strong negative correlation between forebrain acetylcholine (ACH) function and the depth of Alzheimer's dementia (AD). Forebrain norepinephrine (NE) is also frequently reduced in AD. However, it is not clear when these neurochemical abnormalities begin. Since there is some evidence to suggest that the AD begins early in life but fulminates with aging, it may also be the case that ACH and/or NE dysfunction occurs early and participates in the slide towards dementia in old age. To shed light on this possibility, we created lesions of forebrain ACH and NE in the neonatal rat by intracranial injection of 192 IgG saporin and systemic injection of 6-OHDA respectively, allowing the animals to reach old age. Massive ACH and NE lesions were evident at 22 months of age as reflected by immunohistochemical probes for p75 low affinity nerve growth factor and dopamine beta hydroxylase immunoreactive axons respectively. Morris water maze testing showed that surprisingly, the NE but not the ACH lesioned rats were impaired on this reference memory task. Typically, young NE lesioned rats are not impaired on it. The ACH lesion did not exacerbate the consequences of the NE lesion. On the other hand, unbiased stereological counts of hippocampal CA1 pyramidal cells indicated that the ACH lesion caused a significant loss of cells whereas the NE lesion had no effect by itself nor did it exacerbate the effects of the ACH lesion. These results indicate that selective NE and ACH lesions inflicted at birth, persist into old age. Furthermore, the NE lesion seemingly impairs memory in old age while the ACH lesion causes a loss of CA1 cells reminiscent of that which is a hallmark of AD.
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Hypocretin-saporin (Hcrt-sap) as a tool to examine hypocretin function.
Gerashchenko D, Kohls MD, Greco MA, Waleh NS, Salin-Pascual R, Kilduff TS, Lappi DA, Shiromani PJ (2001) Hypocretin-saporin (Hcrt-sap) as a tool to examine hypocretin function. Neuroscience 2001 Abstracts 410.7. Society for Neuroscience, San Diego, CA.
Summary: The hypocretin (Hcrt) peptides are linked to narcolepsy. Humans with narcolepsy have decreased numbers of Hcrt neurons and Hcrt-null mice have narcoleptic symptoms. Hcrt neurons are located only in the lateral hypothalamus (LH) but neither lesions of this nor any other brain region have produced narcoleptic-like sleep, suggesting that specific neurons need to be destroyed. To facilitate lesioning the Hcrt neurons, the ribosome inactivating protein, saporin (SAP), was conjugated to Hcrt-2/orexin B. In vitro binding studies indicated specificity of Hcrt-SAP since it preferentially bound to CHO-cells containing the HcrtR2/OX2 receptor compared to the HcrtR1/OX1 receptor, but not to KNRK cells stably transfected with the NK1 receptor. In vivo specificity was confirmed since administration of the toxin to the LH eliminated some neurons (Hcrt, MCH, and histamine) but not others (alpha-MSH). When the toxin was administered to the LH, rats (n=19) had increased slow wave sleep, REM sleep, and sleep-onset REM sleep periods at night. These were negatively correlated with the loss of Hcrt-containing neurons (r=-0.74; p<0.01). Toxin applied to hypothalamic neurons that are not Hcrt positive but contain the Hcrt receptor lesioned the neurons but did not produce narcoleptic-like sleep. These findings indicate the utility of Hcrt-SAP as a tool and also demonstrate that damage to the LH that also includes a substantial loss of Hcrt neurons is likely to produce the sleep disturbances that occur in narcolepsy.
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Long-term plastic changes in galanin innervation in rat basal forebrain.
Hartonian I, de Lacalle S (2001) Long-term plastic changes in galanin innervation in rat basal forebrain. Neuroscience 2001 Abstracts 254.1. Society for Neuroscience, San Diego, CA.
Summary: Galanin (GAL) immunoreactive (-ir) fibers hyperinnervate remaining cholinergic basal forebrain neurons in Alzheimer's Disease (AD), perhaps exacerbating the cholinergic deficit. The purpose of our study is to determine whether a similar hyperinnervation of GAL-ir fibers occurs following intraparenchymal injection of 192-IgG saporin, a specific cholinergic neurotoxin, within the horizontal diagonal band of Broca (HDB), in 3-month-old rats, and to identify its origin. Immunotoxic lesions produced on average a 31% reduction in cholinergic cell counts on the lesioned side versus the spared side. Hyperinnervation of GAL-ir fibers was observed within and adjacent to the HDB in 28 out of 36 rats, and this effect persisted across time, with 6 months being the longest time examined. Morphometry revealed an increase in the number of GAL-ir cells on the lesioned basal forebrain, as compared to control. A similar change could not be detected in the number of GAL-ir neurons in the amygdala or the bed nucleus of the stria terminalis. Although there was no significant correlation in the amount of cell loss and of GAL hyperinnervation, we suggest that GAL hyperinnervation is triggered by the loss of cells because it is persistent across time. Our data suggests that this hyperinnervation is the result of overexpression of GAL in some cholinergic neurons of the basal forebrain. Since GAL is known to inhibit acetylcholine release, exacerbating the cholinergic dysfunction in AD, this model can be useful to test the efficacy of GAL inhibitors.
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Light- and food-entrained circadian rhythms in rats with neurotoxic lesions of hypocretin cells.
Mistlberger RE, Antle MC, Jones M, Kilduff TS (2001) Light- and food-entrained circadian rhythms in rats with neurotoxic lesions of hypocretin cells. Neuroscience 2001 Abstracts 410.8. Society for Neuroscience, San Diego, CA.
Summary: The hypocretins (Hcrt1 & 2) are lateral hypothalamic (LH) neuropeptides implicated in the regulation of feeding and sleep-wake states. Hcrt lesions attenuate the amplitude of sleep-wake circadian rhythms in rats entrained to light-dark (LD; Gerashchenko et al, Soc. Neurosci Abst., 2000). We examined whether Hcrt cells also participate in the expression of circadian rhythms entrained by daily feeding schedules. Rats received LH injections of Hcrt2 conjugated to the ribosome- inactivating protein saporin. Drinking was recorded in LD 12:12 for 1-2 months, in DD for 48 h, and in LD 2:2 for 24 h, to assess photic entrainment and masking. The rats were then restricted to a 3 h daily meal beginning 6 h after lights-on (LD 12:12). After 29 days, the rats were deprived of food for 50 h. Lesions, assessed by immunocytochemistry using preprohypocretin antibody (Chemicon Int., Inc.), ranged from 0-100% complete. Complete lesions were associated with attenuated mean level and amplitude of circadian drinking rhythms in LD 12:12 and DD, but photic masking in LD 2:2 was unaffected. All rats exhibited food-entrained activity that anticipated feeding time by 1-3 h. This was of lower magnitude in the rat with the largest lesion. These results are consistent with a recent report that Hcrt2-saporin lesions attenuate sleep-wake circadian rhythms, but differ from earlier reports that electrolytic lesions of the LH may potentiate the masking effects of light on behavior. The results suggest that Hcrt cells do not play an essential role in the regulation of circadian rhythms by scheduled feeding.
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Estrogen enhancement of learning requires intact basal forebrain cholinergic neurons.
Gibbs RB (2001) Estrogen enhancement of learning requires intact basal forebrain cholinergic neurons. Neuroscience 2001 Abstracts 312.12. Society for Neuroscience, San Diego, CA.
Summary: We have shown that long-term continuous estrogen replacement, or weekly administration of estrogen plus progesterone, enhances acquisition of a delayed matching-to-position (DMP) spatial memory task. The present study examined whether hormone replacement enhances acquisition of the task after destroying basal forebrain cholinergic neurons with the selective immunotoxin 192IgG-saporin (SAP). Ovariectomized Sprague-Dawley rats received either SAP (1.0 Μg in 1.0 ΜL saline) or vehicle injected directly into the medial septum. Two weeks later, animals received either continuous estrogen replacement (E; 3 mm silastic capsule containing 17-β-estradiol, implanted s.c.) or weekly administration of estradiol (10 Μg in oil s.c.) followed two days later with progesterone (500 Μg in oil s.c.). Controls received sham implantation surgery and vehicle injections. DMP training began after 1 month of treatment. SAP injections significantly impaired acquisition of the DMP task (median days to criterion = 21.0 for SAP-treated animals vs. 14.0 for Ovariectomized controls; p<0.05). Neither continuous E replacement nor weekly administration of E+P significantly enhanced acquisition in the SAP-treated animals. Histology and hippocampal ChAT activity confirmed the loss of cholinergic cells in the medial septum and diagonal band of Broca in SAP treated animals. These findings suggest that cholinergic neurons in the medial septum/diagonal band are necessary for hormone-mediated enhancement of DMP acquisition. Studies using lower doses of SAP are currently underway.
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Hypocretin B-saporin lesions of the brainstem increase rem sleep at night.
Blanco-Centurion CA, Salin-Pascual RJ, Gerashchenko D, Greco MA, Shiromani PJ (2001) Hypocretin B-saporin lesions of the brainstem increase rem sleep at night. Neuroscience 2001 Abstracts 410.9. Society for Neuroscience, San Diego, CA.
Summary: Loss of hypocretin (Hcrt) neurons has been linked to narcolepsy. These neurons project widely throughout brain, but it is not known which projection to which target site produces what symptom of narcolepsy. We (Molec Brain Res, 88:176-182,2001) showed that Hcrt receptors are present in brainstem areas implicated in REM sleep. Since abnormal REM sleep triggering characterizes narcolepsy, we have used Hcrt-saporin, a toxin that selectively lesions Hcrt receptor bearing cells, to assess the effects of such lesions on sleep. In the present study, Sprague Dawley rats (n=21) were administered (under anesthesia) Hcrt-sap (100ng/1ul, vol=0.5 ul bilaterally) or saline to the locus subcoeruleus (LSC) or the medullary inhibitory area of Magoun and Rhines. Subsequently, continuous sleep recordings were made for 21 days. Sleep records were scored blind. In the medulla, Hcrt-sap (n=5) increased the length of REM sleep bouts (p<0.039), which produced a trend towards an increase in REM sleep at night. There were no significant changes in SWS or W. Lesions of the LSC (n=5) increased total sleep time at night (p<0.03) and produced a trend towards a REM sleep increase. Data from both target sites were combined and the Hcrt-sap lesioned rats (n=10 versus saline=11) had a significant increase in REM sleep (30%; p<0.015). At neither site, cataplectic attacks were evident. Our studies with Hcrt-sap indicate site-specific effects on sleep and EEG depending on which Hcrt-receptor bearing neurons are lesioned. All of the symptoms of narcolepsy are evident when the Hcrt-containing neurons are lost. Supported by: NS30140, AG09975, AG15853, MH55772, DVA Med Research.
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Cholinergic lesions of the thalamic reticular nucleus using 192-IgG-saporin.
Tait DS, Latimer M, Woodson W, Brown VJ (2001) Cholinergic lesions of the thalamic reticular nucleus using 192-IgG-saporin. Neuroscience 2001 Abstracts 313.13. Society for Neuroscience, San Diego, CA.
Summary: The TRN is likely to be involved in attention, based on studies of its anatomy, electrophysiological properties and the effects of lesions (see Guillery et al, 1998, TINS 21:28-32). The cholinergic neurons of the basal forebrain (BF) have also been implicated in attention (Sarter and Bruno, 2000, Neuroscience, 95:933-952). Intriguingly, rostral TRN receives cholinergic innervation from the BF (Hallanger et al, 1987, J Comp Neurol, 262:105-124) and yet the role of acetylcholine in the TRN has not been investigated, in part because of difficulty in selectively manipulating the cholineric input to TRN. 192-IgG-saporin is a conjugation of the ribosome-inactivating protein, saporin, with the monoclonal antibody for the p75 neurotrophin receptor (192-IgG). 192-IgG-saporin, injected into BF or cortical BF-terminal regions, can be used to make selective cholinergic BF lesions. The purpose of the study was to investigate whether it would be possible to lesion the BF cholinergic input to TRN, using the immunotoxin 192-IgG-saporin. Male Lister hooded rats (450-500g) were stereotaxically injected with 192-IgG-saporin into TRN. Doses of 1.4, 1.95 or 2.4μg produced lesions of TRN, with a loss of cholinergic cells observed in the BF at all doses. There was also evidence of depletion of cholinergic input to frontal cortex with all doses. At the highest dose, there was cholinergic depletion in hippocampus. These results suggest that 192-IgG-saporin can be used to lesion the TRN. The behavioural effects of these lesions are under investigation.
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Sauro KM, Sweeney M, Saari MJ (2001) Enriched housing, reticular thalamic nucleus and nucleus basalis: Mediators of attention and learning?. Neuroscience 2001 Abstracts 313.14. Society for Neuroscience, San Diego, CA.
Summary: Housing manipulations have been shown to alter behaviour, neural function and morphology. The Reticular Thalamic Nucleus (Rt) may be involved in thalamocortical relay loops possibly acting as a bi-directional filter for sensory information. Similarly, the Nucleus Basalis (NB) may play a role in attention via cholinergic projections to the cortex. In the current experiment 160 female Wistar rats received bilateral lesions of either the Rt (using ibotenic acid), the NB (using 192 IgG-saporin; Advanced Targeting Systems), or both. Following recovery, the rats were allocated to either an enriched or an isolated housing condition for two weeks. Following open field testing, a modification of Tolman's latent learning paradigm was used to evaluate the effects of the treatments on attention. Thus, prior to behavioural testing half of the rats from each group were pre-exposed to the Spatial Orientation Task (SPOT) while the other half were not. Following sacrifice, brain slices were stained for metabolic activity using the cytochrome-oxidase method, for morphology using cresyl violet stain, and for a marker of cholinergic function, acetylcholinesterase. Data analysis revealed significant interactions among the lesions, housing and latent learning and provided some support for the suggestion that the Rt may filter incoming sensory information whereas the NB may mediate behaviourally relevant attention. Lesions and housing effects were confirmed by histology. (Approved by the Animal Care Committee; supported by Nipissing University).
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Visual orienting response and the nucleus basalis of Meynert.
Isaac M, Pearce JM, Muir JL (2001) Visual orienting response and the nucleus basalis of Meynert. Neuroscience 2001 Abstracts 313.2. Society for Neuroscience, San Diego, CA.
Summary: The nucleus basalis of Meynert (nbM), within the basal forebrain, contains a mass of large cholinergic neurones that send axons throughout the cortex. Previous studies have shown the nbM may be involved in attentional processing. The current study attempted to look at the effects of a cholinergic specific nbM lesion (using 192 IgG-saporin) on an attentional task involving two different serial conditioning schedules. In the Consistent condition a light (10s) was followed by a tone (10s), that signaled food. In the Inconsistent condition the light was presented alone for some trials and for other trials the light was followed by the reinforced tone. The orienting response towards the light was measured at three points throughout its presentation: within the first two seconds (light onset) and then twice within the post-onset period (2s-10s). According to the Pearce-Hall theory (1980), animals pay more attention to the light when it is followed by unpredictable events (Inconsistent condition) rather than predictable events (Consistent condition). Accordingly, the orienting response directed towards the post-onset light (2-10s), was stronger for sham operated rats that were trained with the Inconsistent than the Consistent condition. By contrast by the lesioned group was weak and at a similar level for both groups. These results suggest that the nbM lesions prevented enhanced attention to the light in the Inconsistent condition. These findings support the claim that the nbM is important for enhancing attention to stimuli (Chiba et al, 1995).
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Schultz JA, Butt AE, George CL, Garraghty PE (2001) The effects of cortical cholinergic depletion on the performance of adult rats in an appetitive-to-aversive transfer task. Neuroscience 2001 Abstracts 313.3. Society for Neuroscience, San Diego, CA.
Summary: The acetylcholinergic (ACh) projections from the nucleus basalis magnocellularis (NBM) to the neocortex have been implicated in attentional processes. In a test of the hypothesis that only complex learning is affected by damage to this cholinergic system, we examined the effects of NBM lesions in an appetitive-to-aversive transfer learning task. Rats were trained using a tone to signal the availability of food reward for lever-pressing before being transferred to an avoidance learning task where the same tone signaled foot-shock that could be escaped or avoided by lever-pressing. A second experiment examined learning in the aversive context only. For both experiments, male Long-Evans rats received bilateral infusions of the immunotoxin192 IgG saporin into the NBM, sham surgery, or no treatment. Acquisition in the appetitive phase of the appetitive-to-aversive transfer task was normal in the NBM lesion group. However, transfer performance in the aversive task was impaired in NBM lesion group; NBM-lesioned rats acquired the avoidance response more slowly and had lower asymptotic avoidance rates than controls. NBM-lesioned rats tested only in the aversive task performed normally. Thus, the deficit in avoidance learning observed in the NBM-lesioned rats previously trained in the appetitive task was not due simply to an inability to learn in the aversive context. Impairments in transfer learning are instead argued to reflect the relative complexity of the appetitive-to-aversive transfer task as compared to either task alone.
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Yurrita MM, Givens B (2001) Effect of chlordiazepoxide infusions into the basal forebrain on medial prefrontal neural activity of rats during sustained visual attention. Neuroscience 2001 Abstracts 313.4. Society for Neuroscience, San Diego, CA.
Summary: There is extensive evidence suggesting a role for the basal forebrain (BF) cholinergic system in attentional processing. In particular, cortical acetylcholine has been shown to modulate performance in a sustained visual attention task, medial prefrontal cortex (mPFC) neural activity, and distractor-related alterations in mPFC neural activity. In order to further characterize the role of the BF in modulation of attention, the effect of direct infusions of the benzodiazepine receptor agonist chlordiazepoxide (CDP) into the BF was investigated. Specifically, this experiment sought to study the effect of two different doses of CDP (20 and 40µg/hemisphere) on mPFC neural activity of rats performing a task that requires them to discriminate between the presence or absence of short, unpredictable stimuli under testing conditions that vary the level of attentional demand. The overall firing rate of mPFC units recorded during performance in the task was not affected by bilateral infusions of either dose of CDP. There was, however, a differential effect of the two doses on the number of units that show an increase in firing rate during the presentation of the distractor. The high dose of CDP increased the percentage of single units that show a distractor-related increase in firing rate, while the low dose had no effect. In order to determine whether the effect of CDP on mPFC neural activity is mediated via cholinergic projections to cortex, the effect of bilateral CDP infusions into the BF on mPFC neural activity will be studied after local cortical deafferentation of the recording area using 192 IgG-saporin. Supported by: NS37026.
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Camara AL, Pereira EF, Alkondon M, Randall WR, Castro NG, Cintra WM, Albuquerque EX (2001) Septal innervation of the hippocampus regulates expression α7 nicotinic receptors in CA1 and CA3 pyramidal neurons. Neuroscience 2001 Abstracts 145.1. Society for Neuroscience, San Diego, CA.
Summary: To investigate the effects of septal innervation on expression of α7 nicotinic receptors (nAChRs) in CA1 and CA3 pyramidal neurons in the hippocampus, the patch-clamp technique and confocal microscopy were applied to organotypic hippocampal cultures and septal-hippocampal co-cultures. In the co-cultures, septal fibers labeled with DiI were visualized in the hippocampus. Field stimulation of septal fibers also resulted in postsynaptic currents that could be recorded from CA1 and CA3 pyramidal neurons in the hippocampus. These currents had glutamatergic, GABAergic and cholinergic components. The latter originated most likely from the septal cholinergic neurons that were labeled in situ with the cholinergic marker Cy3-192 IgG. α7 nAChRs in the somatodendritic region of CA1 and CA3 pyramidal neurons in the hippocampus in cultures and co-cultures were activated by the α7 nAChR agonist choline, which elicited type IA currents, and were visualized by labeling with rhodamine-conjugated α-bungarotoxin (Rho-α-BGT). After 21 days in vitro, the amplitude of type IA currents was substantially smaller in pyramidal neurons in septal-hippocampal co-cultures than in hippocampal oragnotypic cultures. Labeling of the somatodendritic region of hippocampal pyramidal neurons with Rho-α-BGT was also less intense in the organotypic co-cultures than in cultures. These results suggest that functional septal innervation of the hippocampus regulates the expression of α7 nAChRs in hippocampal pyramidal neurons.
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Nestin expression in neurons of the medial septum/diagonal band in the adult rat.
Yan J, Price DL, Koliatsos VE (2001) Nestin expression in neurons of the medial septum/diagonal band in the adult rat. Neuroscience 2001 Abstracts 25.9. Society for Neuroscience, San Diego, CA.
Summary: Nestin is a marker for neuronal precursor cells in normal animals. In adult animals, nestin (+) cells are limited to the ventricular wall, hippocampus and the rostral migratory stream, where neurogenesis is known to persist throughout life. We are now reporting the existence of nestin (+) cells in the medial septum/diagonal band area based on immunocytochemical staining with different nestin antibodies. Many of these cells colocalize ChAT and nestin. In addition, some nestin (+) cells can be traced with the carbocyanin dye SP-DiI injected into the lateral ventricle to label cell lineages originating in the ependymal layer. Medial septal/diagonal band lesions by complete fimbria-fornix transections or 192-IgG-saporin conjugate injections into the ventricle cause an increase in BrdU (+) and nestin (+) cells in medial septum/diagonal band especially in anterior planes. We are currently double labeling the sections with BrdU and nestin or TUJ1. Our working hypothesis is that there may be ongoing neurogenesis in the medial septum/diagonal band in the adult brain, especially after injury or under pathological conditions and this may have implications for pathogenesis and treatment of Alzheimer's disease.
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Capsaicin-sensitive inhibitory pathway in rat spinal cord dorsal horn.
Gu JG, Nakatsuka T, Tanaka E, Takeda D, Jennifer LX (2001) Capsaicin-sensitive inhibitory pathway in rat spinal cord dorsal horn. Neuroscience 2001 Abstracts 158.13. Society for Neuroscience, San Diego, CA.
Summary: The inhibitory system in the spinal cord plays an important role in regulating nociceptive sensory inputs. Here we examined inhibitory synaptic activity in lamina V neurons of the spinal dorsal horn following the activation of capsaicin VR1 receptors. Experiments were performed with spinal cord slice preparations and inhibitory postsynaptic currents (IPSCs) were recorded using patch-clamp technique. Bath application of capsaicin (2 μM) increased the amplitude and frequency of GABAergic and glycinergic spontaneous IPSCs in the majority of lamina V neurons tested. The effects of capsaicin were completely antagonized by capsazepine (10 μM), and were also blocked in the presence of tetrodotoxin (0.5 μM). However, when CNQX (20 μM) and APV (100 μM) were used to block glutamatergic synaptic transmission, the effects of capsaicin were not abolished. Furthermore, after the injection of IB4-saporin into sciatic nerve to remove IB4-positive C-primary afferent terminals, capsaicin still increased sIPSC frequency in the presence of CNQX and APV. These results suggest that inhibitory pathway could be recruited in the absence of glutamatergic inputs from primary afferents. The release of neuropeptides from capsaicin-sensitive C-primary afferents may activate GABAergic and glycinergic interneurons in superficial laminae, and the inhibitory activity may be further forwarded to lamina V neurons. The capsaicin-sensitive inhibitory pathway may play an important role in the control of nociceptive transmission in the spinal cord.
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Power AE, Thal LJ, McGaugh JL (2001) Memory enhancement induced by post-training norepinephrine in the basolateral amygdala is blocked by 192-IgG saporin lesions of the nucleus basalis magnocellularis. Neuroscience 2001 Abstracts 84.13. Society for Neuroscience, San Diego, CA.
Summary: Drugs and stress hormones act in the basolateral amygdala (BLA) to modulate memory storage. The BLA projects to the nucleus basalis magnocellaris (NBM), which sends broad cholinergic projections to the neocortex. These NBM-cortex projections have been implicated in learning, memory storage and cortical plasticity. The current study was designed to test whether the cholingeric NBM-cortex projections are involved in BLA-mediated memory modulation. Rats were given bilateral cholinergic lesions of the NBM with 192-IgG saporin (0.1 μg/ 0.5 μl per side) or sham infusions, and implanted with bilateral cannulae aimed at the BLA. One week after surgery the rats were trained in the inhibitory avoidance task. Immediately after training, the rats were given bilateral infusions of norepinephrine (0.3μg, 1.0 μg, or 3.0 μg) or vehicle (0.2 μl PBS) into the BLA. On a 48-h retention test, the norepinephrine infusions produced a dose-dependent enhancement of retention (0.3μg and 1.0 μg doses) in sham-operated controls. NBM-lesioned rats that received these memory-enhancing doses of norepinephrine had retention latencies that did not differ from vehicle infused controls. Thus memory enhancement induced by post-training intra-BLA infusion of norepinephrine was blocked in 192-IgG saporin NBM-lesioned rats. ChAT assays of frontal and occipital cortices confirmed the lesions. These findings indicate that the cholinergic NBM-cortex projections are involved in BLA-mediated modulation of memory.
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Nakatsuka T, Takeda D, Gu JG (2001) α,β-methylene ATP sensitive P2X receptor mediated enhancement of glutamate release from the central terminals of Aδ primary afferents onto lamina V neurons in rat spinal cord. Neuroscience 2001 Abstracts 158.16. Society for Neuroscience, San Diego, CA.
Summary: We examined the role of αβmATP-sensitive P2X receptors in modulating glutamate release from sensory synapses of the spinal cord by using whole-cell patch-clamp recordings from dorsal horn neurons in lamina V region. The majority of lamina V neurons synapsed with terminals expressing αβmATP-sensitive P2X receptors. Application of P2X receptor agonist 100 μM αβmATP resulted in a large increase in mEPSC frequency. The increases in mEPSC frequency by αβmATP were completely abolished by the P2X receptor antagonist 10 μM PPADS, but were not blocked by Ca2+ channel blocker 30 μM La3+. αβmATP remained to be effective in increasing mEPSC frequency after the removal of superficial dorsal horn (lamina I-III) or after the injection of IB4-saporin into sciatic nerve to remove P2X3 expressing afferent terminals. Furthermore, we found that αβmATP-sensitive synapses of lamina V neurons were associated with central terminals derived from Aδ primary afferents. The EPSCs evoked by dorsal root stimulation at Aδ-fiber intensity were potentiated by 1 μM αβmATP as well as by the ecto-ATPase inhibitor 10 μM ARL67156, and depressed in the presence of 10 μM PPADS and 5 μM suramin. These results suggest that αβmATP-sensitive P2X receptors play a significant role in modulating excitatory synaptic transmission in the spinal cord.
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Bailey AM, Rudisill ML, Hoof EM, Loving ML (2001) The effects of 192 IgG-saporin lesions to the nucleus basalis magnocellularis/substantia innominata (nBM/SI) on two learning set formation tasks and open field activity. Neuroscience 2001 Abstracts 85.13. Society for Neuroscience, San Diego, CA.
Summary: Male Long Evans rats (Rattus norvegicus) were used to investigate the role of the nucleus basalis magnocellularis/substantia innominata (nBM/SI) in learning set formation. Rats with bilateral 192 IgG-Saporin lesions to the nBM/SI were tested on olfactory discrimination learning set, discrimination reversal learning set, and open field activity. Assessment of open field activity indicated no group differences in general activity levels. Control animals performed significantly better than chance on trial 2 across the 50 problems given in the olfactory discrimination learning set paradigm, suggesting evidence of learning set formation. The nBM/SI lesion group did not perform significantly above chance on trial 2 overall; however, they did perform above chance on trial 2 over the last 10 problems in the olfactory discrimination learning set task. Discrimination reversal followed testing on the olfactory discrimination learning set task. No group differences were seen in discrimination reversal performance. Both control and nBM/SI lesioned animals performed well on the discrimination reversal learning set task, improving with each reversal, and both groups performed significantly higher than expected by chance on trial 2 in the discrimination reversal paradigm, indicating learning set formation. Results suggest that removal of the nBM/SI cholinergic system through 192 IgG-Saporin lesions impairs early acquisition of learning set compared to control animals, but does not interrupt later use of learning set formation.
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Stahl CE, Kusayama T, Keep M, Elmer E, Watanabe S, Borlongan CV (2001) Cyclosporine-A improves performance in passive avoidance task in adult rats with basal forebrain lesions. Neuroscience 2001 Abstracts 101.11. Society for Neuroscience, San Diego, CA.
Summary: While mainly used as an immunosuppressant, newly identified properties of CsA suggest its potential as a therapeutic agent for neurological disorders. In the present study, we investigated the effects of CsA in acquisition and retention of passive avoidance task in adult rats lesioned with 192-IgG-saporin, an immunotoxin that targets cholinergic neurons in the basal forebrain. Starting on the day of the lesion up to 7 days thereafter, animals received either daily CsA (10 mg/kg, i.p. ) or vehicle alone. On day 8 (acquisition), animals were trained in a three-compartment shuttle box passive avoidance task. Twenty-fours later, the retention tests were performed. During the acquisition phase, animals that received CsA treatment significantly entered less in the shock-associated box than those that received vehicle alone. However, the mean total acquisition times between the two groups were not statistically significant. In the retention phase, CsA-treated animals displayed significantly longer latency to stay in the safe compartment compared to vehicle-treated animals. Histological analysis using ChAT immunostaining revealed sparing (80% of control) of basal forebrain cholinergic neurons in CsA-treated animals. The use of CsA may prove beneficial for treatment of neurological disorders characterized by a dysfunctional cholinergic system.
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Works SJ, Wellman CL (2001) Differential dendritic atrophy in frontal cortex after cholinergic lesion in young adult and aged rats. Neuroscience 2001 Abstracts 101.17. Society for Neuroscience, San Diego, CA.
Summary: Previously, we demonstrated that plasticity of frontal cortex is altered in aging rats: three months after surgery, ibotenic acid lesions of the nucleus basalis magnocellularis (NBM) produce larger declines in dendritic morphology in frontal cortex of aged rats relative to young adults. To determine whether the differential effect of the lesion was due specifically to loss of cholinergic input from the NBM, we assessed dendritic morphology in frontal cortex after cholinergic depletion in young adult and aged male rats. Rats received unilateral lesions of the NBM using 192 IgG-saporin, and sham lesions of the contralateral NBM. Two weeks after surgery, brains were stained using a Golgi-Cox procedure. Pyramidal neurons in lamina II-III of frontal cortex were drawn and dendritic morphology was quantified in three dimensions. In young adults, lesions did not alter overall branch number or length. However, in aged rats, lesions decreased basilar dendritic number and length, by 17% and 25% respectively. Furthermore, young adults demonstrated a lesion-induced redistribution of basilar dendrites: dendritic material proximal to the soma was decreased 15%, while distal dendritic material was increased as much as threefold relative to the sham-lesioned hemisphere. Alternatively, lesions in aged rats decreased the amount of distal dendritic material by 25%. Thus, the dendritic atrophy resulting from NBM lesions in aged rats occurs within two weeks after lesion, and results specifically from loss of cholinergic innervation.
Related Products: 192-IgG-SAP (Cat. #IT-01)
Selective cholinergic deafferentation affects GR mRNA expression in the rat brain.
Helm KA, Han JS, Gallagher M (2001) Selective cholinergic deafferentation affects GR mRNA expression in the rat brain. Neuroscience 2001 Abstracts 175.10. Society for Neuroscience, San Diego, CA.
Summary: Two common features of the aging process include progressive dysfunction of both the basal forebrain cholinergic (BFC) system and suprahypothalamic feedback on the hypothalamic-pituitary-adrenal (HPA) axis. Prior research has shown that an age-related reduction in glucocorticoid receptor (GR) mRNA expression occurs in cortical target sites of the BFC system, including the hippocampus, prefrontal cortex, and anterior olfactory cortex, which are highly correlated with both spatial learning impairments and a blunted negative feedback of the stress response among aged rats. Selective deafferentation of the BFC system in young rats produces a similar reduction in both GR mRNA expression in the hippocampus and the efficiency of the stress response, as measured by a protracted increase in the levels of plasma corticosterone following acute stress. The current study investigated the possibility that loss of cholinergic input from cells in the basal forebrain alters GR mRNA expression in other BFC target structures, including the medial prefrontal cortex and the anterior olfactory cortex. Lesions of the BFC system were made by microinjections of the immunotoxin IgG-192-saporin into the medial septum/vertical limb of the diagonal band, and the substantia innominata/nucleus basalis. Basal levels of plasma corticosterone measured in the morning and evening 3 weeks later did not reveal any differences between lesioned and non-lesioned rats. The abundance of GR mRNA in sections processed for quantitative in situ hybridization will include a full analysis of cortical as well as subcortical sites to reveal the extent of effects of cholinergic lesions on GR mRNA expression throughout the brain. Supported by NIA PO1-AG09973.
Related Products: 192-IgG-SAP (Cat. #IT-01)
Schwann cells can enter the demyelinated spinal cord from dorsal roots via scar tissue.
Janni G, Jasmin L, Ohara PT (2001) Schwann cells can enter the demyelinated spinal cord from dorsal roots via scar tissue. Neuroscience 2001 Abstracts 157.4. Society for Neuroscience, San Diego, CA.
Summary: We have studied the routes of entry of Schwann cells into the demyelinated spinal cord. Following application of the toxin CTB-Sap (B fragment of Cholera toxin conjugated to Saporin) into the intrathecal space of adult rats there occurs massive loss of oligodendrocytes with secondary demyelination of the lumbar spinal cord with sparing of axons. Concurrent with the demyelination, an arachnoiditis develops that results in dorsal roots becoming adherent to the lateral spinal cord. Light and electron microscopy showed that Schwann cells in the dorsal roots were always separated from the demyelinated axons by a mesenchymal interface part of which was formed by the dorsal root perineurium. Within 15 days of the demyelination, Schwann cell precursors (p75 immunopositive) migrated from the dorsal roots into the spinal cord via the adhesions and were found to divide within the cord. Schwann cell myelination of demyelinated central axons was consistently observed by day 30. Therefore the Schwann precursors were able to migrate into demyelinated spinal cord through non-neuronal cellular barriers without being in direct contact with demyelinated axons. These findings suggest that, under appropriate conditions, Schwann cells might be introduced therapeutically into the demyelinated spinal cord via intrathecal application and avoid direct spinal injection.
Related Products: CTB-SAP (Cat. #IT-14)
Burk JA, Herzog CD, Porter MC, Mahoney J, Bruno JP, Sarter M (2001) Interactions between partial cortical cholinergic deafferentation and aging on sustained attention performance in rats. Neuroscience 2001 Abstracts 202.5. Society for Neuroscience, San Diego, CA.
Summary: Previous studies have provided only limited support for the idea that aging alone impairs the functions of basal forebrain corticopetal cholinergic neurons. Conversely, aging has been hypothesized to exacerbate the functional consequences of prior insult to, or degenerative processes in, the basal forebrain cholinergic system. The present study assessed the effects of aging on the sustained attention performance of rats with moderate lesion-induced loss of cortical cholinergic inputs. Previous studies on the effects of extensive (> 70 %) cortical cholinergic deafferentation indicated that the integrity of this system is necessary for the animals' ability to detect rare and unpredictable visual signals. In the present longitudinal experiment, moderate (< 50 %) 192 IgG-saporin-induced loss of cortical cholinergic inputs, produced in well-trained, 16 month-old rats, did not – as was intended - immediately affect their attentional performance. Animals continued to undergo attentional performance training for the remainder of their lives. At the age of 31 months, impairments in performance began to emerge in lesioned animals. Compared to their sham-lesioned counterparts, lesioned animals exhibited a steeper decrement in their ability to detect hits in the course of a test session. At this age, the performance of sham-lesioned animals did not differ from their earlier performance prior to and immediately after the lesion. These results support the general hypothesis that aging serves to exacerbate the effects of pre-existing degeneration of the basal forebrain cholinergic system.
Related Products: 192-IgG-SAP (Cat. #IT-01)
Rinaman L, Wonders CP (2001) Targeted destruction of A2/C2 catecholamine neurons alters hypothalamic responses to vagal stimulation. Neuroscience 2001 Abstracts 131.4. Society for Neuroscience, San Diego, CA.
Summary: Central catecholamine (CA) pathways participate in viscerosensory modulation of hypothalamic neuroendocrine function. Different brainstem CA cell groups may relay different types of viscerosensory signals to different classes of hypothalamic effectors. The present study sought to determine the role of dorsal medullary A2/C2 neurons in hypothalamic responses to exogenous cholecystokinin (CCK), which activates gastrointestinal vagal sensory inputs to the caudal brainstem. Saporin toxin conjugated to dopamine-beta-hydroxylase antibody (anti-DbH-sap; 10 ng in 100 nl) or control toxin was microinjected unilaterally or bilaterally into the A2/C2 region of the dorsal vagal complex in adult male rats. After 10-14 days, rats were injected i.p. with CCK (10 ug/kg) and perfused with fixative 1 hr later. Brainstem and forebrain sections were processed for dual immunocytochemical detection of cFos (a marker of neural activation) and DbH (to define the lesion). Additional forebrain sections were processed for cFos and either oxytocin (OT), vasopressin (AVP), or corticotropin-releasing factor (CRF) to identify hypothalamic neurons activated by CCK. Anti-DbH-sap destroyed the majority of A2/C2 neurons within the microinjection site(s), with minimal non-specific damage. A2/C2 lesions markedly attenuated CCK-induced activation of OT neurons and, to a lesser extent, attentuated CRF activation. Conversely, CCK-induced cFos expression was significantly increased in AVP neurons. The latter effect was observed only after bilateral lesions. These results indicate that A2/C2 neurons participate in vagal sensory-mediated stimulation of OT neurons and CRF neurons, and inhibition of AVP neurons.
Related Products: Anti-DBH-SAP (Cat. #IT-03)
Luger NM, Sabino MC, Schwei MJ, Rogers SD, Pomonis JD, Keyser CP, Mach DB, Salak-Johnson J, Clohisy DR, Mantyh PW (2001) Intrathecal infusion of substance P-saporin ablates substance p receptor expressing neurons in the dorsal horn of the spinal cord and attenuates bone cancer pain. Neuroscience 2001 Abstracts 55.4. Society for Neuroscience, San Diego, CA.
Summary: Over 75% of advanced cancer patients must cope with chronic cancer pain. Interestingly, bone cancer pain is the most common and difficult to control. While current therapies are effective in alleviating many aspects of bone cancer pain, they are often accompanied by significant unwanted side effects. To better understand the population of spinal cord neurons that are involved in conveying bone cancer pain and to determine the efficacy of a novel therapeutic modality, we ablated substance P receptor (SPR)+ neurons in the spinal cord using intrathecal infusion of substance P-Saporin (SP-SAP). SP-SAP is a suicide ligand which consists of the ribosomal inactivating factor saporin conjugated to substance P, a peptidergic neurotransmitter involved in nociception. SP-SAP selectively ablates SPR+ neurons located in lamina I and III-V of the spinal cord. C3H male mice received intrathecal SP-SAP treatment 30 days prior to injection of 2472 osteosarcoma cells into the intramedulary space of a femur. Following injection, osteolytic sarcoma cells were confined within the femur by an amalgam plug. Mice were behaviorally tested 17 days post-tumor implantation and both ongoing and movement-evoked pain assessed. Ablation of SPR+ neurons in the dorsal spinal cord coincided with attenuation of both spontaneous and movement-evoked pain behaviors. These results suggest that SPR expressing neurons are involved in the development and progression of the bone cancer pain state and SP-SAP may serve as a useful therapy to treat this debilitating condition. Supported by NIH & VA.
Related Products: SP-SAP (Cat. #IT-07)
Gadd CA, Murtra P, Hall CN, Gana M, Webber MJ, De Felipe C, Hunt SP (2001) NK1-expressing neurons critical for morphine reward behaviors in mice: C-fos expression and ablation of NK1-expressing neurons. Neuroscience 2001 Abstracts 224.13. Society for Neuroscience, San Diego, CA.
Summary: We have previously shown using conditioned place preference (CPP) that mice lacking the preferred receptor for substance P (NK1) show an absence of the rewarding response to morphine as well as reduced conditioned place aversion and physical withdrawal signs following chronic opiate treatment (Nature 405, 180-183). To locate those regions of the brain in which NK1-expressing neurons are crucial for opiate-mediated reward behavior, we examined the expression of c-Fos following acute (10 mg/kg IP) and chronic (increasing doses from 10 to 100 mg/kg IP) morphine administration, and following CPP to morphine (7.5 mg/kg) in wild-type and NK1 knockout mice. The expression of c-Fos in the brains of mice treated with chronic or acute morphine treatment was similar in both genotypes. Moreover, NK1-expressing neurons in the striatum and nucleus accumbens (NAc) were never seen to co-express c-Fos immunoreactivity. In contrast, the expression of c-Fos following the CPP protocol was significantly different between genotypes with a reduced number of c-Fos positive neurons in NK1 knockout mice in the amygdala and hippocampus but not in the NAc or dorsomedial striatum (DMS). We next investigated the effects of selective ablation of NK1 expressing neurons by injecting substance P-saporin into these regions. Our results suggest that destruction of these cells in the amygdala but not in the NAc or DMS causes a reduction in CPP to morphine without affecting anxiety levels or locomotor activity.
Related Products: SP-SAP (Cat. #IT-07)