sfn2002

Guyenet PG, Stornetta RL, Rosin DL, Wang H, Sevigny CP, Weston MC (2002) Somatostatin immunoreactivity is a diagnostic marker of the pre-Boetzinger complex. Neuroscience 2002 Abstracts 362.4. Society for Neuroscience, Orlando, FL.

Summary: Selective ablation of the neurokinin-1 receptor-ir (NK1R-ir) neurons of the ventral respiratory group (VRG) causes major respiratory deficits. Since this population of NK1R-ir neurons is heterogeneous, additional markers are needed to identify which subgroup is most critical to respiratory rhythmogenesis. In the present study, the pre-Boetzinger complex (pre-BoetC) was defined as a 500 μ-long segment of the ventral respiratory group (VRG) located rostral to the spinally projecting inspiratory premotor neurons. This region of the ventral medulla was the only one that contained somatostatin-immunoreactive (SST-ir) neuronal somata. These cells were small (108 μ²), generally fusiform and they expressed very high levels of preprosomatostatin (PPSST) mRNA. All SST-ir neurons were strongly NK1R-ir and were destroyed by saporin conjugated with an NK1R agonist. Most SST-ir neurons (>90%) contained vesicular glutamate transporter 2 (VGLUT2) mRNA whereas <1% contained GAD-67 mRNA and few (6%) contained preproenkephalin mRNA. The results of retrograde labeling experiments with Fluoro-Gold demonstrated that SST-ir neurons do not project to the spinal cord but that over 75% project to the contralateral pre-BoetC. In conclusion, somatic SST immunoreactivity can be used as a diagnostic marker of the pre-BoetC. The SST-ir cells of the pre-BoetC are small glutamatergic interneurons with contralateral projections and they express high levels of NK1 receptors. The homogeneous features of this group of interneurons and their exclusive location in the pre-BoetC suggest that they could be the NK1R-ir neurons whose destruction disrupts respiratory rhythm. (HL 28785 & 60003).

Related Products: SSP-SAP (Cat. #IT-11)

Sugaya K, Qu T (2002) Stem cell transplantation strategies for a lesion model of Alzheimer’s disease. Neuroscience 2002 Abstracts 237.1. Society for Neuroscience, Orlando, FL.

Summary: Stem cell transplantation strategies are advocated in Alzheimer’s disease (AD) neuroregeneration therapy. Since basal cholinergic neurons, which selectively degenerate in AD, extend long projections into the cortex and hippocampus, a stumbling block for neuroreplacement treatment in AD is whether these degenerating cholinergic cells can be replaced by the transplantation of stem cells. To answer this question, we transplanted human neural stem cells (HNSCs) into nucleus basalis magnocelluerlis (NBM) lesion model rats. The lesion was induced either by an injection of ibotenic acid or by anti-NGF receptor antibody conjugated with saporin. HNSCs were labeled by the incorporation of bromodeoxy uridine (BrdU) into the nuclei and simultaneously injected into the contralateral side of the lateral ventricle (Qu, 2001) of the NBM lesioned animal. Four weeks after the surgery, the brain was examined by immunohistochemistry for choline acetyl transferase (ChAT), βIII-tubulin, glial fibrillary acidic protein (GFAP), and BrdU. We detected many GFAP-positive cells in the lesion area, but they were not BrdU-positive, indicating astrocytes activation in this area. We found BrdU-positive cells with ChAT or βIII-tubulin immunoreactivity in the lesion site, indicating that HNSCs migrated to the lesion site and had differentiated into cholinergic and other neuronal cells. These neuronally differentiating HNSCs were rather morphologically premature neurons, and although we have yet to confirm the physiological function or any projections into the hippocampus or cortex, our results could indicate that we have pioneered a positive study of neuroreplacement treatment for cholinergic neurons in AD.

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

Meng ID, Harasawa I, Lai J, Porreca F, Fields HL (2002) Changes in rostral ventromedial medulla (RVM) neurons after the selective loss of mu-opioid receptor expressing cells. Neuroscience 2002 Abstracts 351.9. Society for Neuroscience, Orlando, FL.

Summary: Different subpopulations of RVM neurons inhibit or facilitate dorsal horn nociceptive transmission. Microinjection of saporin conjugated to the mu-opioid receptor (MOR) agonist dermorphin (derm-sap) into the RVM selectively ablates MOR expressing neurons and diminishes neuropathic pain symptoms (Porreca et al., 2001). We examined the properties of neurons surviving a single RVM injection of derm-sap or sap control. Three classes of RVM neurons (On, Off, and Neutral) have been described with distinct responses to noxious stimuli and MOR agonists. On-cells increase and Off-cells cease firing just prior to a tail flick; MOR agonists inhibit On-cells and disinhibit Off-cells. Neutral cells are unaffected by either noxious stimulation or MOR agonists. Using single unit recording in lightly anesthetized rats a total of 10 electrode tracks were made per rat and each unit encountered was characterized according to its tail flick related activity. Injection of derm-sap (n=8) resulted in fewer On- and Off-cells when compared to saporin controls (n=8). The number of Neutral cells remained unchanged. In separate experiments, after derm-sap pretreatment RVM injections of the MOR agonist DAMGO were ineffective whereas injections of the glutamate receptor agonist homocysteic acid into the same sites increased tail flick latencies. The decrease in number of On-cells after derm-sap is consistent with evidence that these neurons express MOR and facilitate nociceptive transmission. The decrease in number of Off-cells indicates that inhibitory neurons responsible for producing the Off-cell tail flick related pause also express MOR.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Herron P, Ismail NS (2002) Effects of cholinergic depletion on the expression of synaptic proteins and functional properties in the rat somatosensory cortex. Neuroscience 2002 Abstracts 256.1. Society for Neuroscience, Orlando, FL.

Summary: Loss of acetylcholine (ACh) has been shown to contribute to numerous cognitive, perceptual, and behavioral deficits in animal studies and in Parkinson and Alzheimer’s patients. The purposes of these experiments were to determine the effects of cholinergic depletion on the expression of glutamic acid decarboxylase (GAD), N-methyl-D-aspartate (NMDA) receptors, synaptophysin, and CaMKII and on functional properties of single neurons in the somatosensory cortex. These experiments were done in the posteromedial barrel subfield (PMBSF) cortex of young adult Sprague-Dawley rats. Selective lesion of cholinergic neurons in the NBM was achieved with intraventricular injections of the immunotoxin (IT), 192 IgG saporin. Electrophysiological recordings and Western blot analyses for the expressions of GAD, NMDA receptors, and synaptophysin were done after a two-week post-injection survival period. The magnitude of evoked and spontaneous activities and the receptive field size of single neurons in the somatosensory cortex were investigated. Recordings and Western blot analyses were obtained from the same area of the PMBSF cortex. Results show that cholinergic depletion causes a significant decrease (11.7%) in the magnitude of evoked activity and an increase (10.7%) in the size of receptive fields. GAD, NMDA receptors, and synaptophysin levels in the in the PMBSF cortex were reduced 25%, 12%, 29%, and 12.5% respectively, in cholinergic depleted animals. Thus, cholinergic depletion leads to effects that significantly alter the expression of synaptic proteins involved in plasticity, learning, and memory.

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

Miller SA, Lappi DA, Wiley RG (2002) Enhanced morphine analgeisa after spinal dermorphin-saporin. Neuroscience 2002 Abstracts 218.1. Society for Neuroscience, Orlando, FL.

Summary: Dermorphin-saporin (derm-sap) is a neuropeptide toxin conjugate which is selective for neurons expressing the mu-opiate receptor (MOR). The dermorphin moiety of the conjugate binds MOR which is then internalized by the neuron, carrying the toxin with it. The saproin moiety inactivates ribosomes resulting in cell death. In the present study we sought to determine the effect of destroying MOR expressing neurons in Lamina II of the spinal cord dorsal horn on baseline thermal pain sensitivity and response to systemic morphine analgesia. 456 ng derm-sap (n=8) and vehicle (n=8) were injected into the lumbar CSF of adult male Sprague Dawley rats using a subarachnoid catheter inserted through the atlanto-occipital membrane and passed cadually to the level of the lumbar enlargement. 10 minutes following toxin injection, the catheters were withdrawn and the animals allowed to recover. When tested on a hotplate at 52C and on tail-flick assay, toxin rats did not differ from rats injected with vehicle. However, the dose-response curves for subcutaneous morphine were significantly shifted to the left (increased potency) in the toxin treated rats when compared with vehicle controls. Histological analysis of multiple dorsal root ganglia failed to reveal evidence of any primary afferent cell loss. We interpreted these findings to indicate that the neurons destroyed by derm-sap are lamina II MOR expressing neurons and play a role in morphine analgesia at high stimulus intensities.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Tait DS, McGaughy JA, Latimer MP, Brown VJ (2002) 192-IgG-saporin lesions of the cholinergic basal forebrain do not impair attentional set-shifting, but do increase latency to dig. Neuroscience 2002 Abstracts 286.2. Society for Neuroscience, Orlando, FL.

Summary: The cholinergic neurons of the basal forebrain which project to cortex, the thalamic reticular nucleus and the amygdala, have been implicated in vigilance and attention (Sarter and Bruno, 2000). This study examined the effects of basal forebrain cholinergic depletion on a shifting of attentional set. Male Lister hooded rats were stereotaxically injected with 192-IgG-saporin into basal forebrain to effect cholinergic depletion. Doses of 0.20mg or 0.25mg resulted in a loss of cholinergic cells in the basal forebrain and depletion of cholinergic input to frontal cortex and the thalamic reticular nucleus. The test of attentional set shifting task for the rat (Birrell and Brown, 2000) measures acquisition, reversal learning and shifting of attention between stimulus dimensions. Trials to criterion and latency to dig were recorded. There was no evidence of impairment in acquisition, reversal learning or set-shifting performance in the rats with cholinergic depletion compared to controls. There was a significant effect on dig latency. This was apparent only when the lesioned rats first approached an incorrect (i.e. unbaited bowl): although no more likely to dig in the incorrect bowl, the lesioned rats took longer to then move to the correct bowl. No effects were seen on dig latency if the rat by chance approached the correct bowl first. We conclude that attentional set-shifting is spared following basal forebrain lesions. Changes in latency in the task might be account for by deficits in sustained attention (attention to task) or related to frustration.

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Kolasa K, Harrell LE (2002) Apoptotic affect of cholinergic denervation and hippocampal sympathetic ingrowth following selective immunolesioning with 192-IgG-saporin in rat hippocampus. Neuroscience 2002 Abstracts 295.16. Society for Neuroscience, Orlando, FL.

Summary: In rat, injection of specific cholinotoxin, 192IgG-saporin, into the medial septum (MS) results not only in a selective denervation of hippocampus(CD), but in an ingrowth of peripheral sympathetic fibers, originating from the superior cervical ganglion, into the hippocampus(HSI). A similar process, in which sympathetic axons invade hippocampus, may also occur in Alzheimer’s disease(AD). Our previous studies using MS electrolytic lesions suggested that HSI and CD appear to induce opposite effect on apoptotic markers. Apoptosis has also been implicated in some aspects of AD. By using 192IgG-saporin we have been able to more specifically and precisely study the affect of apoptosis on HSI and CD. Thus, 12 weeks after injection we measured apoptotic protein expression and DNA degradation using Western blot and in situ techniques e.x. TdT-mediated dUTP nick end labeling(TUNEL). Choline acetyltransferase activity (ChAT) and norepinephrine (NE) level was also detected. Like the previous results, we have found increase in apoptotic markers in CD group, while HSI reduced or normalized apoptotic effect to the control group level. We also found decrease in ChAT activity in HSI and CD groups of dorsal hippocampus.The results of the study suggest that cholinergic denervation is responsible for most of the proapoptotic responses, while hippocampal sympathetic ingrowth produced protective effect in the process of programmed cell death in rat dorsal hippocampus.

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Tavares I, Cobos AR, Almeida A, Lima D (2002) The dyssynaptic pathway from the caudal ventrolateral medulla to the spinal cord is relevant for pain modulation. Neuroscience 2002 Abstracts 351.21. Society for Neuroscience, Orlando, FL.

Summary: The caudal ventrolateral medulla (VLM) exerts α2-adrenoreceptor mediated inhibition of pain transmission at the spinal cord. Anatomical studies described a dysynaptic pathway, connecting the VLM with the spinal cord through the A5 noradrenergic cell group, in which the spinally-projecting A5 noradrenergic neurons give collaterals to the VLM. In order to evaluate the role of the VLM-A5-spinal pathway in pain modulation, retrograde transport of the neurotoxin saporin-anti-dopamine-β-hydroxylase (SAP-anti-DBH) from the VLM was used. The VLM of Wistar rats was injected with 0.5μl of a 1% SAP-anti-DBH solution or saline (control group). Four days later, all animals were injected with 50 μl of 5% formalin in the ipsilateral hindpaw, and pain behavior and noxious-evoked spinal c-fos expression, were evaluated. In the SAP-anti-DBH group, a 27% decrease in DBH-immunoreactive neuronal population at the A5 noradrenergic cell group was detected and neuronal death was confirmed by Fluojade staining. Hyperalgesia was detected in the second phase of the formalin test. The numbers of Fos-immunoreacted neurons in the spinal dorsal horn increased. The data suggest that the VLM-A5-spinal pathway participates in pain modulation. It remains to be ascertained whether the lack of effect at the first-phase of the formalin test is due to an insufficient destruction of the A5 noradrenergic cell group or whether it points to a differential effect of this pathway in the two phases of the formalin test.

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Helm KA, Ziegler DR, Gallagher M (2002) Habituation and dexamethasone (DEX) suppression of the stress response following selective lesions of cholinergic input to hippocampus in rats. Neuroscience 2002 Abstracts 370.1. Society for Neuroscience, Orlando, FL.

Summary: Hippocampal neurons have been identified as targets for glucocorticoids that exert inhibitory control over hypothalamic-pituitary-adrenocortical (HPA) axis activity. Prior research has shown that selective removal of cholinergic input to the hippocampus reduces mRNA expression for low-affinity glucocorticoid receptors, while leaving unaffected both mineralocorticoid receptor mRNA and basal levels of circulating corticosterone (CORT). The current study investigated the possibility that loss of cholinergic support from cells in the basal forebrain alters the CORT response to stress. Cholinergic lesions were made by microinjections of the immunotoxin 192 IgG-saporin into the medial septum/vertical limb of the diagonal band, and 3 weeks later rats were subjected to six daily sessions of 30 min restraint stress. Blood samples taken before, during and after stress on Day 1 revealed a prolonged elevation of CORT in response to acute stress in cholinergic lesioned rats. After 5 days of chronic stress, however, both groups significantly habituated to the stressor, as indicated by similarly low CORT profiles throughout both the response and recovery period. Against this similar background, rats were administered a Dexamethasone (DEX) challenge on Day 6, and DEX-induced suppression of endogenous CORT before, during and after stress was attenuated in lesioned rats. These results indicate a mechanism whereby loss of cholinergic function (e.g. in aging and Alzheimer’s Dementia) may compromise the dynamic range of sensitivity to glucocorticoid mediated stress pathways in the brain.

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Ohara PT, Boudah A, Jasmin L (2002) Long-term effects on pain behavior of decreased spinal noradrenaline in neuropathic rats. Neuroscience 2002 Abstracts 351.22. Society for Neuroscience, Orlando, FL.

Summary: We sought to determine if a permanent reduction in the noradrenergic (NA) input to the spinal cord in adult rats would alter the pain behavior associated with nerve injury. Selective NA denervation of the lumbo-sacral cord was achieved by intrathecal injection of anti-dopamine beta-hydroxylase antibodies conjugated to the toxin saporin in 12 female rats. Spinal NA denervation was confirmed histologically in all animals. Saline injected rats served as controls. Two weeks after toxin or saline treatment, a unilateral peripheral neuropathy was induced by tight ligation of the left L5 spinal nerve in both groups. Unexpectedly, the same degree of mechanical hyperalgesia was present in the neuropathic paw of rats in both the toxin and saline treated groups. Rats lacking NA spinal afferents, however, were less responsive to the antinoiceptive effects of morphine administered systemically or intracerebroventricularly. Also, toxin treated rats did not display opioid dependant stress analgesia. Finally, toxin treated rats were more responsive to the antinociceptive effect of the NK1 antagonist CP 96,345 but not to its enantiomer CP 96,344. From these results we conclude that the permanent loss of spinal NA does not alter neuropathic pain behavior, possibly because of compensatory changes in the CNS. The decreased response to opioids is consistent with the previous suggestions of an interaction between noradrenergic and opioidergic systems in producing analgesia. The increased response to NK1 antagonists shows that NA tonically inhibits substance.

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