Search PubMed⌕ Search

Biomedical subjects

R G Wiley

Publications and source records attributed to R G Wiley.

At least 37 records · Page 2Linked to original sources
PubMed1998

Selective lesion of the cholinergic basal forebrain causes a loss of cortical neuropeptide Y and somatostatin neurons.

Degeneration of the cholinergic basal forebrain (CBF) and changes in cortical neuropeptide levels have been reported in Alzheimer's disease. In the present study, we sought to determine if a selective cholinergic lesion of nucleus basalis magnocellularis (Nbm) could affect the number and distribution of neuropeptide Y (NPY) and somatostatin (SS) immunoreactive neurons in the frontoparietal and occipital cortices of rats. Brain sections were evaluated at survival times of 1, 2, 4, 8, 12, 24, 48, 78 and 100 weeks after intraventricular injection of 192-saporin, an immunotoxin directed at the low affinity neurotrophin receptor (p75NGFr), that selectively destroys the CBF. Following the immunotoxin lesion of the Nbm, the number of NPY-labeled neurons decreased 33% in the frontoparietal cortex and 60% in the occipital cortex compared to age-matched normal controls at most survival time points. A significant loss of SS-labeled neurons in both cortical regions was seen 12 weeks after 192-saporin injection with no further change up to 100-week survival time. The effect of age on neuropeptidergic populations was evaluated in normal control rats. The number of NPY and SS immunoreactive neurons in aged rats (21-26 months) decreased by 42% in the frontoparietal cortex and 27% in the occipital cortex when compared with young (3-6 months) and middle-age (9-14 months) rats. When both non-lesioned and lesioned animals with different ages were pooled for linear regression, a significant correlation was found between the number of cortical NPY- and SS-labeled neurons and cortical acetylcholinesterase (AChE) histochemical staining intensity. These findings indicate that: (1) cholinergic denervation of the Nbm is associated with an irreversible loss of neocortical NPY and SS immunoreactive neurons analogous to that observed in Alzheimer's disease and aging; (2) the degree of the loss of cortical NPY and SS immunoreactive neurons seems to be related to the extent of the reduction of cortical AChE intensity in both toxin-injected and normal aged rats. These findings may reflect a trophic dependence of NPY and SS neurons on cortical cholinergic input.

Acetylcholinesterase↗
PubMed1998

Nerve gas-induced seizures: role of acetylcholine in the rapid induction of Fos and glial fibrillary acidic protein in piriform cortex.

Soman (pinacolymethylphosphonofluoridate), a highly potent irreversible inhibitor of acetylcholinesterase (AChE), causes seizures and rapidly increases Fos and glial fibrillary acidic protein (GFAP) staining in piriform cortex (PC). This suggests that the inhibition of AChE by soman leads to increased acetylcholine (ACh) and neuronal excitability in PC. The sole source of cholinergic input to PC is from the nucleus of the diagonal band (NDB). To investigate the role of ACh in soman-induced seizures, we lesioned cholinergic neurons in NDB unilaterally with 192-IgG-saporin. By 10 d, saporin eliminated staining for choline acetyltransferase (ChAT), the synthetic enzyme for ACh, in NDB ipsilateral to the lesion. Staining for AChE, the degradative enzyme for ACh, was eliminated in PC ipsilateral to the lesioned NDB. By 45-60 min after soman, increased Fos and GFAP staining in PC was evident only ipsilateral to the unlesioned NDB. By 90-120 min after soman, Fos and GFAP staining increased bilaterally in PC. In a second experiment, electrical stimulation electrodes were implanted unilaterally in the NDB to activate focally the projections to PC in unanesthetized rats. Within 5 min of NDB stimulation, there were clear behavioral and EEG signs of convulsions. After 45-60 min of NDB stimulation, there was increased Fos and GFAP staining in layer II of PC ipsilateral to the stimulation site. Pretreatment with the selective muscarinic receptor antagonist scopolamine blocked the convulsions and prevented increased Fos and GFAP staining in PC. These results suggest that ACh release in PC triggers the initiation of seizures and gliosis after soman administration, predominantly by the activation of muscarinic receptors.

Acetylcholine↗
PubMed1998

Destruction of locus coeruleus neuronal perikarya after injection of anti-dopamine-B-hydroxylase immunotoxin into the olfactory bulb of the rat.

Saporin, a ribosome-inactivating protein, was coupled to a monoclonal antibody to dopamine-B-hydroxylase (DBH) and injected unilaterally into the olfactory bulb of rats. After 4-13 days survival, the rat brain was processed histologically and the locus coerulei (LC) examined with Nissl and anti-DBH staining. There were degenerating dendrites in surviving LC neurons on the side ipsilateral to the immunotoxin-injected olfactory bulb. The number of Nissl-positive LC neurons in a transverse section through the caudal one third of the LC was reduced from 116+/-10 to 50+/-8 neurons (P < 0.01, n = 7) and the number of DBH-positive neurons in the more rostral LC sections was reduced from 13+/-2 to 5+/-1 (P < 0.05, n = 4). Our results indicate that it is possible to lesion LC neurons via retrograde intraaxonal transport of saporin-anti-DBH immunotoxin from the olfactory bulb.

Animals↗
PubMed1998

Lesion-induced transneuronal plasticity of the cholinergic innervation in the adult rat entorhinal cortex.

The present experiments were designed to determine the effect that lesions of the basal forebrain cholinergic system exert on cholinergic interneurons within the entorhinal cortex (EC) in the rat. Unilateral infusion of 192 IgG-saporin into the nucleus of the horizontal diagonal band of Broca (HDB) decreased the number of ipsilateral choline acetyltransferase immunoreactive (ChAT-ir) neurons by 54%. Two-four weeks after the lesion, the ipsilateral EC exhibited a moderate but significant loss of ChAT-ir fibres and interneurons. Adjacent sections revealed a parallel loss of vasoactive intestinal polypeptide (VIP) immunoreactivity. Cell counts in the cingulate cortex were unaffected, suggesting that this effect was indeed specific to the main target area for HDB neurons. Ibotenic acid lesions also induced a significant 36% decrease in the number of cholinergic neurons in the ipsilateral HDB, and disappearance of ChAT terminals in the EC, whereas the number of ChAT-ir neurons in the EC was unchanged. Since ibotenic acid affects all cells and not only cholinergic ones, our results suggest that the specific degeneration of cholinergic neurons in the HDB after 192 IgG-saporin treatment could be inducing transsynaptic effects on their targets. Injections of 192 IgG-saporin directly into the EC also lesioned the cholinergic projection from the HDB, but had no effect on the intrinsic population. Eight weeks after immunolesion, the number of interneurons immunoreactive for ChAT and VIP in the EC had returned to normal values, and persisted for as long as 6 months after the lesion. By contrast, ChAT-ir neurons in the HDB were permanently lost. Our results suggest that the transient down-regulation of the cholinergic phenotype in entorhinal cortex interneurons could be a manifestation of activity-dependent plasticity, and that the loss of cholinergic innervation from the basal forebrain could be responsible for these effects through an imbalance of inputs. We hypothesize that the recovery of the phenotypic expression of entorhinal interneurons could be due to a recovery in their innervation, perhaps from sprouting axons in the same fields, belonging to surviving cholinergic neurons in the basal forebrain.

Animals↗
PubMed1998

Coordinate expression of the vesicular acetylcholine transporter and choline acetyltransferase following septohippocampal pathway lesions.

The gene for the vesicular acetylcholine transporter (VAChT) was recently cloned and found to be located within a 5' noncoding intron of the gene for choline acetyltransferase (ChAT). There appear to be several shared and unique promoters for each gene, suggesting that control of expression of these two genes can be either coordinated or independent. Two lesions, axotomy and immunotoxin, directed at the well defined septohippocampal cholinergic pathway were used to determine VAChT and ChAT protein expression in the degenerating terminal fields in the hippocampus and the cell bodies of the medial septum nucleus after injury. Two weeks after lesioning, decreases of up to 90% in VAChT were found in the affected hippocampus by immunoblotting and immunocytochemistry, similar to ChAT activity. The number of VAChT- and ChAT-immunopositive neurons in the medial septum decreased by up to 95%. Eight weeks following axotomy, the number of VAChT- and ChAT-immunopositive neurons had increased to almost 50% in fimbria-fornix-lesioned animals, indicating coordinate reexpression of both cholinergic markers in recovered neurons. There was no recovery of either VAChT or ChAT immunoreactivity after the irreversible immunotoxin lesions. Thus, with use of immunological techniques, there appears to be coordinate expression of VAChT and ChAT in the septohippocampal pathway following either unilateral fimbria-fornix or bilateral immunotoxin lesion.

Animals↗
PubMed1998

Neonatal treatment with 192 IgG-saporin produces long-term forebrain cholinergic deficits and reduces dendritic branching and spine density of neocortical pyramidal neurons.

The role of basal forebrain-derived cholinergic afferents in the development of neocortex was studied in postnatal rats. Newborn rat pups received intraventricular injections of 192 IgG-saporin. Following survival periods ranging from 2 days to 6 months, the brains were processed to document the cholinergic lesion and to examine morphological consequences. Immunocytochemistry for choline acetyltransferase (ChAT) and in situ hybridization for ChAT mRNA demonstrate a loss of approximately 75% of the cholinergic neurons in the medial septum and nucleus of the diagonal band of Broca in the basal forebrain. In situ hybridization for glutamic acid decarboxylase mRNA reveals no loss of basal forebrain GABAergic neurons. Acetylcholinesterase histochemistry demonstrates a marked reduction of the cholinergic axons in neocortex. Cholinergic axons are reduced throughout the cortical layers; this reduction is more marked in medial than in lateral cortical areas. The thickness of neocortex is reduced by approximately 10%. Retrograde labeling of layer V cortico-collicular pyramidal cells reveals a reduction in cell body size and also a reduction in numbers of branches of apical dendrites. Spine densities on apical dendrites are reduced by approximately 20-25% in 192 IgG-saporin-treated cases; no change was detected in number of spines on basal dendrites. These results indicate a developmental or maintenance role for cholinergic afferents to cerebral cortical neurons.

Acetylcholinesterase↗
PubMed1998

Role of the basal forebrain cholinergic projection in somatosensory cortical plasticity.

Trimming all but two whiskers in adult rats produces a predictable change in cortical cell-evoked responses characterized by increased responsiveness to the two intact whiskers and decreased responsiveness to the trimmed whiskers. This type of synaptic plasticity in rat somatic sensory cortex, called "whisker pairing plasticity," first appears in cells above and below the layer IV barrels. These are also the cortical layers that receive the densest cholinergic inputs from the nucleus basalis. The present study assesses whether the cholinergic inputs to cortex have a role in regulating whisker pairing plasticity. To do this, cholinergic basal forebrain fibers were eliminated using an immunotoxin specific for these fibers. A monoclonal antibody to the low-affinity nerve growth factor receptor 192 IgG, conjugated to the cytotoxin saporin, was injected into cortex to eliminate cholinergic fibers in the barrel field. The immunotoxin reduces acetylcholine esterase (AChE)-positive fibers in S1 cortex by >90% by 3 wk after injection. Sham-depleted animals in which either saporin alone or saporin unconjugated to 192 IgG is injected into the cortex produces no decrease in AChE-positive fibers in cortex. Sham-depleted animals show the expected plasticity in barrel column neurons. In contrast, no plasticity develops in the ACh-depleted, 7-day whisker-paired animals. These results support the conclusion that the basal forebrain cholinergic projection to cortex is an important facilitator of synaptic plasticity in mature cortex.

Acetylcholine↗
PubMed1997

Inhibition of hyperalgesia by ablation of lamina I spinal neurons expressing the substance P receptor.

Substance P is released in the spinal cord in response to painful stimuli, but its role in nociceptive signaling remains unclear. When a conjugate of substance P and the ribosome-inactivating protein saporin was infused into the spinal cord, it was internalized and cytotoxic to lamina I spinal cord neurons that express the substance P receptor. This treatment left responses to mild noxious stimuli unchanged, but markedly attenuated responses to highly noxious stimuli and mechanical and thermal hyperalgesia. Thus, lamina I spinal cord neurons that express the substance P receptor play a pivotal role in the transmission of highly noxious stimuli and the maintenance of hyperalgesia.

Animals↗
PubMed1997

Selective changes in hippocampal neuropeptide Y neurons following removal of the cholinergic septal inputs.

The number and distribution of subpopulations of hilar interneurons containing neuropeptide Y (NPY), somatostatin (SOM), or gamma-aminobutyric acid (GABA) immunoreactivities were examined in the hilus of the dentate gyrus following removal of the cholinergic septal inputs. One, 2, 4, 8, 12, and 24 weeks after intracerebroventricular injections of immunotoxin, consisting of antibody to the low-affinity nerve growth factor receptor conjugated to saporin (192 IgG-saporin), lesioned rats were processed simultaneously with controls for NPY, SOM, or GABA immunolabeling. Across all time points, the number of NPY-labeled neurons was reduced to a statistically significant level (paired t-test, P = 0.001) in the injected rats (73% of control values, on average). The decrease in the number of NPY-labeled neurons was not limited to any particular subregion rostrally but appeared greater in the central region caudally. The size of NPY-labeled neurons did not differ statistically between control and immunolesioned rats examined at 1, 2, and 24 week time points. In contrast, the number of both SOM- and GABA-immunoreactive neurons in injected rats did not appear to be affected in any consistent manner. Examination of the hilus in adjacent Nissl-stained sections with the optical dissector revealed that although the total number of small nonprincipal cells (5-15 microm in diameter) did not appear affected at the 4-week time point, there was a statistically significant (P = 0.03) reduction across the 8-24-week time points (to 80% of control values, on average). Dual-labeling studies on separate rats showed that a small subpopulation of the NPY- and SOM-labeled neurons, primarily in the infragranular hilus, were colocalized with neurons containing GABA immunoreactivity (18% and 5%, respectively). These studies demonstrate that removal of the cholinergic septal inputs (1) can cause relatively rapid, selective decreases in the number of NPY-immunoreactive hippocampal interneurons and (2) appears to lead to the death of hippocampal interneurons over a longer time course. The changes in NPY immunoreactivity seem to occur in the portion of interneurons that probably does not contain either SOM or GABA immunoreactivity.

Animals↗
PubMed1997

Destruction of neurokinin-1 receptor expressing cells in vitro and in vivo using substance P-saporin in rats.

Substance P (SP) acts on neurons through the neurokinin-1 (NK-1) receptor. Conjugation of SP to the ribosome inactivating protein, saporin (SAP), produces a cytotoxin selective for cells that express the NK-1 receptor. SP-SAP cytotoxicity was inhibited by pre-treating the toxin to reduce the disulfide bond connecting SP to SAP or by pre-incubation with anti-SP antiserum or by SP analog showing that SP-SAP acts through binding of the SP moiety to NK-1 receptors. Injection of SP-SAP into the striatum selectively destroyed NK-1 receptor expressing interneurons. These results show that SP-SAP will be useful for studying the function of NK-1 receptor expressing neurons.

Animals↗
PubMed1997

Selective ablation of rat knee joint innervation with injected immunotoxin: a potential new model for the study of neuropathic arthritis.

Neuropathic arthropathy is a severe chronic degenerative condition associated with decreased or absent sensory innervation of the involved joint. Existing animal models of neuropathic arthritis are limited by the technical difficulties of obtaining either highly selective or complete joint denervation in a minimally invasive fashion. We undertook experiments to determine the feasibility of using the newly described method of selective neuronal lesioning with injected immunotoxin as a means of creating a more tractable model of neuropathic arthritis. Retrograde tracing with fluorochrome revealed that the knee joint of the female Wistar rat is supplied by 581 +/- 31 (mean +/- SD) joint afferents. Immunohistochemistry confirmed that virtually all sensory neurons in the rat express the cell surface receptor Thy 1. Injection of rat knee joints with an immunotoxin targeted toward Thy 1 resulted in the selective ablation of an average of 88% of the joint afferents identified with fluorochrome that are normally found in the ipsilateral L3 and L4 ganglia.

Afferent Pathways↗
PubMed1997

The effects of selective cholinergic basal forebrain lesions and aging upon expectancy in the rat.

The effects of selective cholinergic cell loss within the basal forebrain (BF) were determined using a task that requires shifting of attention between two visual stimuli. Discriminability between two stimuli and response bias were determined in young and old F-344 rats given BF injections of IgG-192 saporin (100 ng). The lesion reduced ChAT activity in the frontal and parietal cortices, hippocampus, and olfactory bulbs. The lesion did not significantly alter Na+/K(+)-ATPase activity in cortex, hippocampus, or olfactory bulbs, or endogenous levels of neuropeptide Y and neurokinin B within the BF. The BF lesions impaired both stimulus discriminability and response bias in young and old rats. The BF lesions had a significantly greater effect upon stimulus discriminability and response bias in aged rats, compared to young rats, only when the stimulus duration was very brief, i.e., when the task was most difficult to solve. At longer stimulus durations, aging and lesions showed no interaction. The results suggest that the selective loss of cholinergic cells in the BF, but not normal aging, impairs the ability to discriminate between independent sensory stimuli. The loss of these cells confers a response bias in simple operant tasks involving motor responses to reward-related visual stimuli.

Acetylcholine↗
PubMed1997

Retrograde degeneration and colchicine protection of basal forebrain cholinergic neurons following hippocampal injections of an immunotoxin against the P75 nerve growth factor receptor.

Intracerebroventricular injection of 192 IgG antibody against the p75LNGFR rat low affinity nerve growth factor receptor conjugated with saporin, a ribosome inactivating protein, has been shown to destroy the p75LNGFR-expressing cholinergic neurons of the basal forebrain. We injected this immunotoxin into the hippocampus and studied its retrograde effect upon the cholinergic neurons of the medial septum and the vertical limb of the diagonal band of Broca. Seven days after injection, there was a nearly total depletion of cholinergic axons within the hippocampus. This depletion was associated with a marked and significant decrease in the number of cholinergic neurons of the ipsilateral medial septum and the vertical limb of the diagonal band of Broca. At longer survival times, these changes were more pronounced. Parvalbumin-positive, GABAergic neurons within the same areas of the basal forebrain were not affected by immunotoxin injections. Injections of saporin alone had no effect upon cholinergic neurons. Simultaneous injection of colchicine with the immunotoxin resulted in a significant reduction of retrograde degeneration of cholinergic neurons and relative preservation of hippocampal cholinergic axons. These observations suggest that 192 IgG-saporin is transported retrogradely from the hippocampus to the cholinergic neurons in the medial septum and the vertical limb of the diagonal band of Broca and provide a model for retrograde degeneration of basal forebrain cholinergic neurons following cortically based toxic-pathologic processes.

Animals↗
PubMed1996

Central noradrenergic lesioning using anti-DBH-saporin: anatomical findings.

The ability to create lesions of discrete neuronal populations is an important strategy for clarifying the function of these populations. The power of this approach is critically dependent upon the selectivity of the experimental lesioning technique. Anti-neuronal immunotoxins offer an efficient way to produce highly specific neural lesions. Two previous immunotoxins have been shown to be effective in both the CNS and PNS. They are OX7-saporin, which is targeted at Thy1, and 192-saporin, which is targeted at the low affinity neurotrophin receptor, p75NTR. In the present study, we sought to determine if an immunotoxin targeted at the neurotransmitter synthesizing enzyme, dopamine beta-hydroxylase (DBH), could selectively destroy central noradrenergic neurons after intraventricular administration. This immunotoxin, which consists of a monoclonal antibody to DBH coupled by a disulfide bond to saporin (a ribosome inactivating protein), has been shown to be selectively toxic to peripheral noradrenergic sympathetic neurons in rats after systemic injection. In the present study, immunohistochemical and Cresyl violet staining showed that the noradrenergic neurons of the locus coeruleus are destroyed bilaterally after intraventricular (i.c.v.) injection of 5, 10, and 20 micrograms of anti-DBH-saporin (alpha-DBH-sap) into rats. Complete bilateral lesioning of the A5 and A7 cell groups occurred at the two higher doses. Lesions of the A1/C1 and A2/C2/C3 cell groups were incomplete at all three doses. Dopaminergic neurons of the substantia nigra and ventral tegmental area and serotonergic neurons of the raphé, all monoaminergic neurons that do not express DBH, survived all alpha-DBH-sap doses. The cholinergic neurons of the basal forebrain, which are selectively killed by i.c.v. injection of 192-saporin, and cerebellar Purkinje cells which are killed by OX7-saporin, were not killed by alpha-DBH-sap. These results show that alpha-DBH-sap efficiently and selectively destroys CNS noradrenergic neurons after i.c.v. injection. The preferential destruction of locus coeruleus, A5, and A7 over A1/C1 and A2/C2/C3 may be due to more efficient access of the immunotoxin to these neurons and their terminals after i.c.v. injection.

Adrenergic Fibers↗
PubMed1996

192 IgG-saporin lesion of basal forebrain cholinergic neurons in neonatal rats.

Seven day old rats received bilateral intraventricular injections (200 ng) of the immunotoxin 192 IgG-saporin. When assayed in adulthood, these rats showed an 84% loss of hippocampal and a 52% loss of cortical choline acetyltransferase (ChAT) activity. ChAT was unaffected in the caudate. Cholinergic neurons immunoreactive (IR) for the low affinity neurotrophin receptor (P75NTR) were severely reduced throughout the basal forebrain nuclei. Cortical and hippocampal norepinephrine were increased and these areas showed ingrowth of ectopic, P75NTR and dopamine beta-hydroxylase IR varicosities. These were probably sympathetic axons. No obvious forebrain dysmorphogenesis was observed and cortical thickness was unaffected. These rats showed no evidence of impaired spatial learning/memory as assessed by the Morris water maze and delayed spatial alternation. However, they were less active on the elevated plus apparatus and spent less time on the open arms, suggestive of increased timidity. 192 IgG-saporin appears to be a powerful tool to selectively lesion basal forebrain cholinergic neurons in the neonatal rat. Surprisingly, the neuromorphological and behavioral sequelae seem minimal. It may be necessary to achieve near-total neonatal destruction of forebrain cholinergic neurons before severe, lasting mnemonic effects are evident.

Acetylcholine↗
PubMed1996

Extensive reinnervation of the hippocampus by embryonic basal forebrain cholinergic neurons grafted into the septum of neonatal rats with selective cholinergic lesions.

Reconstruction of the septohippocampal pathways by axons extending from embryonic cholinergic neuroblasts grafted into the neuron-depleted septum has been explored in the neonatal rat by using a novel lesioning and grafting protocol. Neonatal ablation of the basal forebrain cholinergic projection neurons, accompanied by extensive bilateral cholinergic denervation of the hippocampus and neocortex, was produced at postnatal day (PD) 4 by 192 immunoglobulin (IgG)-saporin intraventricularly. Four days later, cholinergic neuroblasts (from embryonic day 14 rats) were implanted bilaterally into the neuron-depleted septum by using a microtransplantation approach. The results show that homotopically implanted septal neurons survive and integrate well into the developing septal area, extending axons caudally along the myelinated fimbria-fornix and supracallosal pathways that are able to reach the appropriate targets in the denervated hippocampus and cingulate cortex as early as 4 weeks postgrafting. Moreover, the laminar innervation patterns established by the graft-derived axons closely resembled the normal ones and remained essentially unchanged up to at least 6 months, which was the longest postoperative time studied. The reinnervating fibers restored tissue choline acetyltransferase activity (up to 50% of normal) in the dorsal hippocampus and the parietooccipital cortex. Retrograde labeling with Fluoro-Gold from the host hippocampus combined with immunocytochemistry confirmed that most of the projecting neurons, indeed, were cholinergic. The results suggest that the graft-host interactions that are necessary for target-directed axon growth are present in the septohippocampal system during early postnatal maturation. Thus, the present approach may contribute to overcome the functional limitations inherent in the use of ectopically placed intrahippocampal transplants.

Acetylcholine↗
PubMed1996

Expression of m1 and m4 muscarinic receptor mRNA in the striatum following a selective lesion of striatonigral neurons.

Cholinergic stimulation has opposing effects on striatopallidal and striatonigral neurons. Most striatal projection neurons express m1 muscarinic receptor mRNA with m4 mRNA found in 40-50%. Expression of m4 mRNA is found in most preprotachykinin neurons but only a subset of preproenkephalin neurons, suggesting preferential localization of m4 receptors to striatonigral neurons. A volkensin lesion of striatonigral neurons reduced striatal m4 mRNA by 63% and m1 mRNA by only 18%, suggesting that preferential expression of m4 receptors by striatonigral neurons may contribute to their differential response.

Animals↗