sfn2002

Gallegos RA, Lee RS, Crawford E, Wills DN, Carr JR, Zhukov VI, Slaght KE, Huitron-Resendiz S, Criado JR, Henriksen SJ (2002) Selective lesion of ventral tegmental area neurons expressing mu-opioid receptors alters EEG power spectrum across sleep/wake cycle. Neuroscience 2002 Abstracts 276.14. Society for Neuroscience, Orlando, FL.

Summary: The ventral tegmental area (VTA) has long been implicated in motivated behaviors. Our previous study (Lee et al, J Neurosci 2001) also suggests a role for VTA GABAergic neurotransmission in REM sleep. In the current study the potential role of the VTA in modulating electroencephalogram (EEG) activation was explored by selectively deactivating mu-opioid receptor expressing cells in the VTA. Under sodium pentobarbital anesthesia, rats received either (1) a sham operation (2) a single bilateral VTA injection of NMDA (3) a saporin injection or (4) an injection of a dermorphin-saporin (DERM-SAP) conjugate (Advanced Targeting Systems, San Diego). Animals were also fitted with skull electrodes for recording the EEG. The filtered EEG was recorded continuously for 24 hours beginning 21 to 28 days after surgery. Frequency analysis of the EEG in 15-sec epochs revealed differences in the distribution of relative power in the DERM-SAP animals, compared to controls. Low frequency components (0.5-3.0 Hz and 3.0-8.0 Hz) were enhanced in dual lesioned animals during the dark phase but only during sleep. These results indicate that a selective inactivation of cells in the VTA has specific effects on arousal mechanisms in the intact animal.

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

Burgess SE, Gardell LR, Xie Y, Ossipov MH, Vanderah TW, Malan TP, Porreca F, Lai J (2002) Dermorphin-saporin targets descending facilitation in the rostral ventromedial medulla (rvm) to block CCK-induced abnormal pain. Neuroscience 2002 Abstracts 351.11. Society for Neuroscience, Orlando, FL.

Summary: Abnormal pain from L5/L6 spinal nerve ligation (SNL) has been shown to require a time-dependent activation of descending facilitatory pathways arising in the RVM. Additionally, RVM microinjection of L365,260, a cholecystokinin (CCKB) receptor antagonist, reverses SNL-induced tactile and thermal hyperalgesia. These observations suggest the possibility that RVM CCK might “drive” such facilitation from the RVM by activating the endogenous descending facilitation system. Rats were treated with a single RVM injection of dermorphin (DERM) (μ opioid agonist), unconjugated saporin (SAP), or dermorphin-saporin (DERM-SAP) and responses to non-noxious tactile (von Frey filaments) or noxious radiant heat stimuli applied to the hindpaw were measured before and after RVM microinjection of CCK to uninjured rats. RVM DERM-SAP, DERM or SAP did not significantly alter baseline sensory thresholds over 28 days post-injection. At day 28, the rats received bilateral microinjections of CCK (30ng) in the RVM. Rats pretreated with DERM or SAP showed a time-related and revsersible CCK-induced tactile and thermal hypersensitivity. In contrast, RVM CCK failed to produce changes in sensory threshold in animals pretreated with DERM-SAP. The RVM pretreatments did not alter responses in control rats challenged with CCK vehicle. Additionally, lesions of the dorsolateral funiculus also blocked RVM CCK-induced tactile and thermal hypersensitivity. These data support the possibility of CCK-mediated activation of descending facilitation from the RVM as a mechanism of neuropathic pain.

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

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.

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

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.

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

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.

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