sfn2001

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.

Related Products: Anti-DBH-SAP (Cat. #IT-03)

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.

Related Products: 192-IgG-SAP (Cat. #IT-01)

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.

Related Products: 192-IgG-SAP (Cat. #IT-01)

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.

Related Products: 192-IgG-SAP (Cat. #IT-01)

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.

Related Products: SP-SAP (Cat. #IT-07)

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)