targeted-toxins

Seki S, Erickson KA, Sasaki K, Sugaya K, Chancellor MB, de Groat WC, Yoshimura N (2002) Effect of intrathecal substance P-saporin conjugate on bladder hyperactivity induced by intravesical capsaicin. Neuroscience 2002 Abstracts 68.2. Society for Neuroscience, Orlando, FL.

Summary: Substance P and neurokinin 1 (NK1) receptors in the spinal cord reportedly play an important role in the micturition reflex as well as in nociceptive responses. We investigated the effect of elimination of NK 1 receptor-expressing spinal cord neurons using the substance P-saporin conjugate (SSP-SAP) on the micturition reflex. Using female rats, an intrathecal catheter was implanted at the level of the L6-S1 spinal cord for injection of either 8 µl of saporin (1.5 µM) or SSP-SAP (1.0 µM). Awake cystometry performed 3 weeks after injection showed no changes in normal bladder function in saporin and SSP-SAP-treated rats. When bladder hyperactivity was induced by intravesical instillation of capsaicin (15 µM), the reduction of intercontraction interval (ICI) was significantly smaller (43.0 ± 6.2% of reduction) in SSP-SAP-treated rats than in saporin-treated rats (59.3 ± 3.1%). Immunohistochemical staining revealed that the area positively stained with NK1 receptor antibodies in the lamina I of the dorsal horn was significantly reduced by 34% in SSP-SAP treated rats, compared with saporin-treated rats. These results suggest that NK1 receptor-expressing neurons in the dorsal horn of the spinal cord play an important role in inducing bladder hyperactivity elicited by intravesical capsaicin. Thus elimination of NK1 receptor-expressing neurons in the spinal cord using SSP-SAP at this concentration could be effective to treat bladder hyperactivity induced by bladder irritation without affecting normal bladder function.

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Chang Q, Gold PE (2002) Residual hippocampal cholinergic functions after 192 IgG-saporin lesions of the medial septum/ventral diagonal band (MS/VDB). Neuroscience 2002 Abstracts 82.8. Society for Neuroscience, Orlando, FL.

Summary: 192 IgG-saporin (SAP) injections into the MS/VDB reportedly destroy acetylcholine (ACh) neurons projecting to hippocampus (HC) without associated impairments of learning and memory on HC-dependent tasks. These findings contrast sharply with those obtained with many other methods showing close associations between ACh functions in the HC and learning and memory. The present experiment addressed this conflict. SAP was injected into MS/VDB 1 week before the start of neural and behavioral measures. We found: 1) Using in vivo microdialysis, release of ACh in the HC was not abolished but was ~30% of control values. The percent increase in ACh release during spontaneous alternation testing was greater in SAP-treated than in control rats. 2) SAP-treated rats had significant impairments on the HC-dependent alternation task. Moreover, intra-HC injections of physostigmine (20 ng in 1 µl), an indirect ACh agonist, enhanced alternation scores. 3) Physostigmine (0.6 mg/kg, IP) induced more intense tremors in SAP-treated than in control rats. 4) While SAP-treated rats had near-total depletion of cells in MS/VDB stained using immunocytochemistry for choline acetyltransferase (ChAT), quantitative densitometry showed no depletion of ChAT staining in either CA1 or the dentate gyrus in the SAP group. These findings suggest that there are residual and even heightened ACh functions which can compensate for the insult of MS/VDB SAP lesions, complicating the use of this lesion to evaluate the role of septohippocampal cholinergic projections in learning, memory or other functions.

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

Truitt WA, McKenna KE, Coolen LM (2002) Lesions of spinothalamic neurons in lumbar spinal cord disrupt ejaculatory reflexes in male rats. Neuroscience 2002 Abstracts 69.2. Society for Neuroscience, Orlando, FL.

Summary: Previously we tested the significance of a population of lumbar spinothalamic (LUST) cells for male sexual behavior in rats. Anatomically, LUST cells are positioned to relay ejaculation-related sensory signals from reproductive organs to the brain and express substance P receptors as well as several neuropeptides including galanin. Ablation of LUST neurons by the selective toxin SSP-saporin resulted in a complete disruption of ejaculatory behavior. These results suggested that LUST cells play a pivotal role in generation of ejaculatory behavior and may be part of a spinal ejaculation generator. To test this hypothesis, we investigated ejaculatory reflexes in male rats with LUST lesions, using the urethrogenital reflex model. SSP-saporin (4 ng/µl) was injected bilaterally into L3-L4 region in sexually experienced male Sprague Dawley rats. Ten days following surgery, animals were deeply anesthetized and spinal cords were transected at upper thoracic levels. Next, urethral stimulation was provided and muscle contractions were recorded in the bulbocavernous muscle (BCM). Following the experiment, animals were sacrificed and lesions were confirmed using immunostaining for galanin, a marker for LUST cells. In non-lesioned animals (n=5), urethral stimulation produced stereotypical reflex contraction of the BCM, and penile reflexes were observed. In contrast, in animals with complete lesions of LUST cells (n=5) the urogenital reflex was severely attenuated. These results indicate that LUST cells are involved in control of ejaculatory reflexes and are part of a spinal ejaculation generator. Supported by: NIH R01 MH60781(LMC)

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Ricceri L, Moles A, Pezzola A, Popoli P, Calamandrei G (2002) Neonatal basal forebrain cholinergic lesions disrupt retention of socially transmitted food preferences and alter EEG activity in adult rats. Neuroscience 2002 Abstracts 82.9. Society for Neuroscience, Orlando, FL.

Summary: Previous studies using selective neonatal lesions of basal forebrain cholinergic neurons showed mild long-term effects on spatial discrimination capabilities, whereas water maze learning appeared intact. In the present study we examined long-term effects of icv injections of 192 IgG saporin performed in 7-day-old rats on the social transmission of food preferences (a form of non-spatial associative memory) at adulthood. In 6-month-old rats the neonatal cholinergic lesion impaired 4-h and 24-h retention of a learned social food preference relative to controls, despite performance on an immediate retention trial was indistinguishable from controls. A second experiment excluded alterations in neophobia towards unfamiliar scented food after neonatal cholinergic lesions: level of novel food consumption did not differ between neonatally saporin-lesioned and control rats. Computerized EEG spectral analysis (FFT transform) performed in 6-month-old rats revealed that the neonatal cholinergic lesions increased δ power and reduced β power in both fronto-parietal and parieto-occipital cortex. Effectiveness of the neonatal lesion was confirmed by a marked cholinergic loss in both hippocampal and cortical regions. Altogether, behavioral and electrophysiological data suggest that the neonatal cholinergic lesion of the basal forebrain – more than the adult one – could represent a useful experimental model of Alzheimer-like memory dysfunctions.

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Mattsson A, Lindqvist E, Ögren SO, Olson L (2002) Amphetamine hyperresponses in cholinergically denervated rats and alpha7 nAChR knockout mice, and effects of nicotinic agonists. Neuroscience 2002 Abstracts 136.7. Society for Neuroscience, Orlando, FL.

Summary: We have recently shown that cholinergic denervation of the basal forebrain in adult rats, using intracerebroventricular injections of the cholinergic immunotoxin 192-saporin, leads to overreactivity in dopaminergic systems, in the form of enhanced amphetamine-induced hyperactivity. This increased sensitivity to amphetamine in cholinergically denervated rats can be partially counteracted by nicotine if given before the amphetamine challenge. The results provide a possible link between deficits in central cholinergic systems and overactivity in dopaminergic systems, which might be of relevance for the pathogenesis of schizophrenia. To further evaluate the role of cholinergic dysfunction for dopaminergic hyperactivity we have focused on the α7 nAChR due to its presumed involvement in schizophrenia. First, we tested if a selective α7 nAChR agonist, AR-R17779, could mimic the effect of nicotine (partial blockade of amphetamine hyperresponse) in cholinergically denervated rats. Second, we tested if mice lacking the α7 nAChR gene have a similar increased sensitivity to amphetamine, as cholinergically denervated rats. Preliminary results from these studies show that AR-R17779 normalizes the effect of amphetamine cholinergically denervated rats, and further, that lack of the α7 nAChR gene leads to an increased sensitivity to amphetamine. Cholinergic deficiencies that cause dopaminergic overactivity are compatible with a role of the α7 nAChR in schizophrenia and may suggest a role for the cholinergic system in other psychotic states.

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Dinh TT, Duffy P, Ritter S (2002) Immunotoxin lesion of spinally projecting catecholamine neurons impairs the adrenal medullary response to glucoprivation and the sympathetic response to forced swim. Neuroscience 2002 Abstracts 76.5. Society for Neuroscience, Orlando, FL.

Summary: Distinct populations of hindbrain catecholamine neurons project spinally to innervate sympathetic and adrenal medullary preganglionic neurons. Previously we injected the immunotoxin, saporin conjugated to anti-dopamine beta hydroxylase (DSAP), into the spinal cord to selectively lesion these neurons. DSAP lesions abolished adrenal medullary Fos expression following insulin-induced hypoglycemia or 2-deoxy-D-glucose (2DG) and eliminated the hyperglycemic response to 2DG, which is mediated by adrenal medullary epinephrine (E) secretion. Here we examine the plasma E and norepinephrine (NE) responses to 2DG (250 mg/kg, s.c.) and to 5 min of forced swim in rats injected at T2-T4 with DSAP or unconjugated saporin (SAP) control solution. Blood was sampled remotely via jugular catheters between 0 and 240 min after 2DG or swim. Immunohistochemistry confirmed loss of dopamine B-hydroxylase throughout the spinal cord of DSAP rats. In DSAPs, both plasma E and hyperglycemic responses to 2DG were abolished or severely impaired compared to SAPs. 2DG did not elevate plasma NE in either group. Swim stress increased NE in both SAPs and DSAPs, but the DSAP response was only 60% of the SAP response. Results show for the first time that the selective activation of the adrenal medulla by glucoprivation, described previously, is mediated by spinally projecting catecholamine neurons. Results also demonstrate that spinal catecholamine terminals, presumeably arising from different hindbrain neurons, contribute to, but are not entirely responsible for, sympathetic neuronal responses to swim stress.

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Varga C, Grosche J, Brauer K, Seeger J, Harkany T, Hartig W (2002) Cholinergic neurons in the rabbit forebrain: Chemoarchitecture, in vivo labeling, immunolesions. Neuroscience 2002 Abstracts 35.3. Society for Neuroscience, Orlando, FL.

Summary: While the rabbit basal forebrain and its cholinergic components became useful targets for modeling of neuropathological changes associated with Alzheimer’s disease, their neuroanatomical organization is still largely elusive. Hence, we focused on (i) the number of cholinergic basal forebrain neurons (CBFN)in the major nuclei based on choline acetyltransferase (ChAT) immunoperoxidase labeling, (ii) the density of ChAT-immunoreactive fibers in distinct neocortical and hippocampal areas, (iii) mapping of projecting CBFN by low-affinity neurotrophin receptor p75 (p75NTR ) staining and (iv) the double fluorescence labeling of ChAT and the neuronal markers p75NTR, nitric oxide synthase (NOS), calbindin, calretinin, parvalbumin, tyrosine hydroxylase and substance P. While cholinergic interneurons were found in the hippocampus, they were not detectable in the neocortex. CBFN were shown to abundantly co-express p75NTR, except in the substantia innominata and ventral pallidum. Whereas cholinergic neurons were devoid of most investigated markers, a subset also contained calbindin or NOS. The selective in vivo labeling of CBFN was achieved with intracerebroventricularly (i.c.v.) injected carbocyanine 3-conjugated ME20.4IgG that recognizes an extracellular epitope of p75NTR. Parallel experiments revealed that the i.c.v. injection of ME20.4IgG-saporin conjugates led to the specific immunolesion of cholinergic cells in about one week, whereas long-term effects of the immunotoxin remain to be further elucidated.

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Turchi JN, Saunders RC, Mishkin M (2002) Effects of cholinergic deafferentation of rhinal cortex on visual recognition in monkeys. Neuroscience 2002 Abstracts 82.5. Society for Neuroscience, Orlando, FL.

Summary: Excitotoxic lesions of the rhinal (perirhinal/entorhinal) cortices yield substantial deficits in visual recognition (Baxter and Murray, 2001; Malkova et al., 2001). To evaluate the mnemonic role of cholinergic inputs to this region, we compared the visual recognition performance of untreated monkeys with that of monkeys given rhinal cortex infusions of the selective cholinergic immunotoxin ME20.4-SAP. This toxin binds to the p75 receptor, borne by corticopetal cholinergic neurons of the basal forebrain, and is retrogradely transported to the cell body where it permanently destroys ribosomal function. Both groups were first trained to criterion in the rule for delayed nonmatching-to-sample (DNMS) with trial-unique stimuli at a 10-s delay in a Wisconsin General Testing Apparatus. This was followed by treatment and recovery for the experimental group (n=3) and an equivalent rest period for the control group (n=4), after which both groups were retrained on the DNMS rule and then given a memory performance test with increasing delays (30, 60, and 120 s) and list lengths (3, 5, 10, and 20 stimuli). The experimental group relearned the DNMS rule without significant impairment but then demonstrated robust deficits when tested with increasing delays (a mean of 83% vs 95% for controls) and list lengths (67% vs 86% for controls). The findings complement results obtained in a study of muscarinic receptor blockade in the perirhinal cortex (Tang et al., 1997) and indicate that cholinergic integrity of the rhinal cortex is critical for visual recognition memory.

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Mohapel P, Leanza G, Lindvall O (2002) Alterations in forebrain acetylcholine influence hippocampal neurogenesis in the adult rodent. Neuroscience 2002 Abstracts 23.9. Society for Neuroscience, Orlando, FL.

Summary: Little is known about how various experiential, environmental and pathological factors regulate neurogenesis in the adult hippocampus. Since the hippocampus receives abundant cholinergic innervation and contains some of the densest distributions of acetylcholine (ACh) fibers, we investigated its potential role in adult neurogenesis. Adult rats received multiple bromodeoxyuridine (BrdU) injections 3 weeks following lesions of the adult rodent forebrain cholinergic projections by intracerebroventricular infusions of 192 IgG-saporin. The day following BrdU administration we observed a significant 20% to 30 % decrease in proliferation in the subgranular cell layer of the dentate gyrus with ACh lesioning. This decrease persisted through to 4 weeks after BrdU administration, when most proliferated cells co-expressed neuronal markers. Conversely, in a separate experiment, naive rats receiving simultaneous injections of the ACh agonist physostigmine and BrdU demonstrated a 30 % increase in proliferated cells (1 day later) and neurons (4 weeks later) in the subgranular cell layer. Our data indicate that cholinergic mechanisms in the forebrain are involved in the regulation of neurogenesis and that this effect may be indirect or direct in the hippocampus.

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

Suzuki R, Morcuende S, Webber M, Hunt SP, Dickenson AH (2002) Superficial NK1-expressing neurons control spinal excitability through activation of descending pathways. Nat Neurosci 5(12):1319-1326. doi: 10.1038/nn966

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