sfn2011

Roland JJ, Stewart AL, Savage LM, Servatius RJ, Pang KCH (2011) Reduced hippocampal acetylcholine efflux after medial septal-diagonal band (MSDB) GABAergic lesion is associated with impaired working memory: behavioral and neurochemical effects of physostigmine. Neuroscience 2011 Abstracts 513.10. Society for Neuroscience, Washington, DC.

Summary: The medial septum provides cholinergic innervation of the hippocampus and changes in hippocampal acetylcholine (ACh) have been tied to memory; deficits and enhancements in memory are correlated with decreases or increases of ACh, respectively. Damage of GABAergic MSDB neurons impaired spatial working memory in a delayed non-match to position task with a 30-s retention interval (DNMTP). Interestingly, lesions reduced maze activated hippocampal ACh efflux, but did not alter basal hippocampal ACh efflux. The current study has two aims. First, is performance impaired and ACh efflux reduced in a non-match to position task (NMTP) with a 0-s retention interval following GABAergic MSDB damage? Second, is performance on DNMTP improved by enhancing hippocampal ACh efflux? Male Sprague-Dawley rats received intraseptal PBS or GAT1-saporin (to damage GABAergic neurons) and a ventral hippocampal microdialysis cannula to assess ACh efflux. In Exp. 1, rats were trained on NMTP for 10 days and received microdialysis on either day 2 (early) or day 9 (late). GAT1-saporin rats were not behaviorally impaired and hippocampal ACh efflux was similar in both treatment groups. These results suggest that performance with a short retention interval (NMTP) is more independent of MSDB influences than training with a long retention interval (DNMTP). Exp. 2 was designed to determine whether the reduced ACh efflux is a critical factor in impaired DNMTP performance in rats with GABAergic MSDB damage. In Exp. 2, all rats will receive 10 day of DNMTP training. On days 8 and 9, rats will be administered (i.c.v.) either saline or the acetylcholinesterase inhibitor, physostigmine (5μg/μl). The effects of physostigmine on behavioral performance and hippocampal ACh efflux will be determined. We predict that physostigmine will increase ACh efflux but not improve behavior, suggesting that hippocampal ACh is not important for DNMTP performance. However, support that both MSDB cholinergic and GABAergic neurons are important for DNMTP performance would be seen if physostigmine increases ACh efflux and enhances DNMTP performance. In summary, damage of MSDB GABAergic neurons modulates hippocampal ACh efflux during performance of a working memory task. Whether hippocampal ACh release plays a critical role in impaired working memory will be answered by these studies.

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Winter SS, Köppen JR, Stout JM, Cameron HA, Wallace DG, Cheatwood JL (2011) Growth factor infusion increases BrdU-positive cells in the denervated medial septum following 192-IgG-saporin lesion. Neuroscience 2011 Abstracts 331.04. Society for Neuroscience, Washington, DC.

Summary: During the progression of Alzheimer’s Disease, degeneration of basal forebrain structures is associated with a decline in mnemonic function and frequently results in episodes of wandering behavior. Previous work has demonstrated that the septohippocampal cholinergic system uniquely contributes to rat spatial orientation. Enhancement of endogenous adult neurogenesis represents one potential method to restore function to the septohippocampal system. Therefore, we tested the hypothesis that co-infusion of epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) would increase the number of new cells in the medial septum following a lesion of the cholinergic system produced by focal injection of the ribosome-inactivating selective immunotoxin 192-IgG-saporin in rats. For this, rats received injections of 192-IgG-saporin into the medial septum. At the same time, a cannula was placed in the lateral ventricle and attached to a subcutaneously-placed osmotic minipump containing either 1) EGF, bFGF, and bromodeoxyuridine (BrdU), or 2) BrdU alone. Infusion of growth factors and BrdU continued for a period of two weeks, at which point the pumps were removed. At 21 days following 192-IgG-saporin injury, rats were perfused following standard protocols. Cryostat sections were collected at 40 microns and were processed via double-fluorescent immunochemistry (IHC) using antibodies against BrdU and doublecortin (DCX). Photomicrographs of BrdU and DCX immunofluorescence were captured under epifluorescence and the number of BrdU-positive and DCX-positive cells was quantified. We detected significantly higher numbers of BrdU-positive cells in the medial septum of rats that received growth factors compared to rats that received BrdU-only (p<0.05). These results indicate that infusion of growth factors following 192-IgG-saporin lesion of the medial septum resulted in an increase in the number of new immature neurons in the medial septum. Studies aimed at determining the fate of these young neurons and their influences on spatial orientation are ongoing.

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Taxini CL, Bícego K, Takakura A, Moreira T, Gargaglioni L (2011) Noradrenergic neurons of the A5 region play a role on hypoxic ventilatory response in unanesthetized rats. Neuroscience 2011 Abstracts 345.09. Society for Neuroscience, Washington, DC.

Summary: The ventrolateral pons contains the A5 group of noradrenergic neurons which is involved in cardiorespiratory control. These cells are strongly activated by carotid body stimulation and display central respiratory modulation. Recently, we showed that A5 neurons contribute to the cardiorespiratory effects elicited by chemoreflex stimulation in anesthetized rats. In the present study, we assessed the role of A5 noradrenergic neurons on cardiorespiratory responses produce by hypoxia in unanesthetized rats. To selectively destroy noradrenergic neurons, we administered the immunotoxin anti-dopamine β-hydroxylase-saporin (anti-DβH-SAP, 200nL) bilaterally in the A5 region of male Wistar rats (n = 8). Hypoxia (7% O2, 30 min) produced an increase in ventilation (Ve) (1470 ± 141 mLkg-1min-1), respiratory frequency (RF) (179 ± 139 brethsmin-1) and heart rate (484 ± 29 bpm), without affect mean arterial pressure (MAP) in conscious rats. Bilateral destruction of the catecholaminergic A5 neurons reduced the hypoxia-induced hyperventilation (942 ± 110 mLkg-1min-1, p<0.05), increase in RF (139 ± 11 breathsmin-1, p<0.05) and tachycardia (399 ± 39 bpm, p<0.05). These results suggest that A5 noradrenergic neurons contribute to the increase in heart rate, ventilation and respiratory frequency during peripheral chemoreflex stimulation.

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Donahue RR, Corder GF, Mcnamara KC, Wiley RG, Taylor BK (2011) Transmission of neuropathic pain by spinal neurons expressing the NPY Y1 receptor. Neuroscience 2011 Abstracts 179.16. Society for Neuroscience, Washington, DC.

Summary: Endogenous neuropeptide Y (NPY) acts at Y receptors in the dorsal spinal cord to exert a tonic inhibitory control of chronic allodynia (Solway et al, PNAS 108:7224-9, 2011). In this and the adjacent presentation, we tested the hypothesis that NPY does this by inhibiting Y1 receptors on pain transmission neurons or on central terminals of primary afferent neurons. We selectively lesioned cells expressing the NPY receptors in the dorsal horn with intrathecal administration of the NPY-conjugated ribosomal toxin, NPY-saporin. NPY-saporin significantly reduced the population of Y1 receptors in the lumbar dorsal horn by over 50%. Neither NK1 receptors in the dorsal horn, nor neuronal counts in the DRG were affected, suggesting a specific effect on Y1+, NK1- neurons in the dorsal horn, while sparing Y1+ central presynaptic terminals. Fourteen days later, we ligated the tibial and common peroneal branches of the sciatic nerve (spared nerve injury, SNI), and evaluated the development of allodynia and hyperalgesia on post-SNI days 1, 3, 5, 7, 14, 21, 28, 35, and 42. When compared to saporin controls, NPY-saporin (1000 ng) decreased mechanical allodynia (von Frey threshold), cold allodynia (paw withdrawal response to application of a drop of acetone) and mechanical hyperalgesia (paw response to blunt pin). This effect began three days after SNI and lasted until forty two days after SNI. When injected in uninjured rats, NPY-saporin did not disrupt motor coordination (accelerating rotarod), baseline heat or mechanical thresholds, or animal activity levels. We conclude that Y1-expressing cells in the dorsal horn exert a tonic facilitatory control of neuropathic pain, and partially mediate the inhibitory actions of NPY.

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Corder GF, Donahue R, Winter MK, Chen W, Mccarson KE, Marvizon J, Taylor B (2011) Activation of postsynaptic NPY Y1 and presynaptic Y2 receptors reduce spinal nociceptive transmission. Neuroscience 2011 Abstracts 179.17. Society for Neuroscience, Washington, DC.

Summary: Exogenous (Intondi et al, Neuroscience, 2008) and endogenous (Solway et al, PNAS 108:7224-9, 2011) neuropeptide Y (NPY) acts at Y1 and Y2 receptors in the dorsal horn (DH) to inhibit hypersensitivity to mechanical and thermal stimuli. The adjacent poster (Donahue, et al, SFN 2011) describes our use of a targeted NPY-saporin neurotoxin approach to selectively remove spinal cord (SC) neurons expressing the Y1 receptor — the data implicate a contribution of Y1-expressing, pain transmission neurons to behavioral signs of persistent pain. To determine whether persistent noxious input is associated with a compensatory increase in NPY-mediated inhibitory signaling (presumably at Y1-expressing DH neurons), we performed GTPγS binding assays in SC slices taken from animals following the intraplantar (i.pl) injection of complete Freund’s adjuvant (CFA). CFA significantly reduced the EC50 of Y1 agonist (Leu31,Pro34-NPY)-induced [35S]GTPγS binding in ipsilateral DH to 0.24 ± 0.17 μM, as compared to sham (1.38 ± 0.51 μM). This support the hypothesis that injury increases in the efficiency of coupling between Y1-receptors and G-proteins. To determine whether compensatory NPY inhibition occurs at presynaptic sites, we studied the activity of presynaptic Y2 receptor in NPY-saporin-treated rats. Intrathecal injection of the Y2 receptor antagonist BIIE0246 reduced von Frey thresholds (saporin group from 1.3±0.4 to 0.6 ±0.1g; 750 ng NPY-saporin group from 5.4±1.0 to 1.2±0.2g, p<0.05), suggesting that presynaptic Y2 receptors contribute to a tonic endogenous inhibition of inflammatory pain. In support of this hypothesis, BIIE0246-induced hyperalgesia (21 days after CFA) significantly increased the Emax of Y2 agonist (PYY3-36)-induced [35S]GTPγS binding. We next determined whether NPY acts at presynaptic terminals of primary afferent neurons to reduce the release of substance P (SP). First, in both the i.pl carrageenan and CFA models of inflammatory pain, intrathecal administration of NPY reduced in vivo neurokin-1 (NK1) receptor internalization (an indirect measure of functional SP release). Second, application of either (Leu31,Pro34)-NPY) or PYY3-36 to spinal cord slices concentration-dependently reduced NK1 internalization in the ipsilateral dorsal horn evoked by electrical stimulation of the dorsal root (1000 pulses of 20 V, 0.4 ms at 100 Hz); these effects were reversed by the Y1 antagonist BIBO3304. We conclude that injury up-regulates post-synaptic Y1 and pre-synaptic Y2 spinal inhibitory mechanisms to reduce behavioral signs of persistent pain.

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Sinha SP, Roland JJ, Servatius RJ, Pang KCH (2011) The role of medial septal/diagonal band GABAergic neurons in proactive interference: Effects of selective immunotoxic lesions in latent inhibition. Neuroscience 2011 Abstracts 199.22. Society for Neuroscience, Washington, DC.

Summary: The medial septum/diagonal band (MSDB) is a critical structure for learning and memory, yet the functional contributions of its individual neuronal populations (including cholinergic, GABAergic, glutamatergic and peptidergic cells) are still being characterized. Recent studies have implicated a contributing role for the GABAergic MSDB neuronal population, as selective immunotoxic GABAergic lesions of the MSDB (with GAT1-saporin) produce behavioral impairments in spatial and instrumental tasks. Compared to intact controls, rats with GABAergic MSDB lesions are impaired in learning new spatial locations in a delayed match to position procedure and also exhibit a slower rate of extinguishing a previously acquired avoidance response – behaviors that are consistent with an exacerbation of proactive interference. To further establish the role of these neurons in proactive interference, this study examined the effects of selective GABAergic MSDB lesions in latent inhibition (LI) of the classically conditioned eyeblink response. LI in delay eyeblink conditioning is a phenomenon in which pre-exposure to the conditioned stimulus (CS) interferes with the subjects’ ability to subsequently associate the CS with an unconditioned stimulus (US), resulting in slower acquisition of the conditioned response (CR). We hypothesized that if damage of GABAergic MSDB neurons increases proactive interference, then rats with selective lesions of these neurons would show facilitated LI. Male Sprague-Dawley rats (n=18) were administered either phosphate-buffered saline or GAT1-saporin via intracranial injection into the MSDB. After 7-10 days of recovery, electrodes were implanted into the upper eyelids of the rats for delivery of US and EMG recording. Conditioning began after another 5-7 days of recovery, with Day 1 consisting of 30 minutes of acclimation to the conditioning context. Day 2 began with either 30 presentations of the CS (82dB, 500ms white noise, 25 – 35s ITI) or context pre-exposure of equal duration, followed immediately by 100 paired CS-US trials (82 dB, 500ms white noise co-terminating with a 10V, 10ms square-wave stimulus). In preliminary results, intraseptal GAT1-saporin did not alter CR acquisition in context pre-exposed rats. Rats with GABAergic MSDB lesions continued to exhibit latent inhibition. These preliminary results do not support the idea that damage of GABAergic MSDB neurons increase proactive interference of the classically conditioned eyeblink response. Future studies will examine whether manipulations of the number of CS pre-exposures would facilitate LI in rats with GABAergic MSDB lesions.

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Van Der List DA, Chapman B (2011) Targeted ablation of intrinsically photosensitive melanopsin expressing retinal ganglion cells early in development alters retinal morphology within the inner plexiform layer of mice. Neuroscience 2011 Abstracts 232.12. Society for Neuroscience, Washington, DC.

Summary: It has been demonstrated in adult mice, that eliminating a small subset of retinal ganglion cells expressing the photopigment melanopsin (ip-RGCs) with an immunotoxin alters the effects of light on circadian rhythms. The immunotoxin was made by conjugating the melanopsin antibody with ribosome-inactivating protein, saporin. It has also been observed that the ablation of ip-RGCs in adult mice did not alter retinal morphology. Specifically, it was found that dendrites arising from starburst amacrine cells retained their position within the inner plexiform layer (IPL) suggesting no reorganization within this synaptic layer (Goz et al. 2008). In this study, we used the same melanopsin immunotoxin (Mx) (Advanced Targeting Systems) to perform intravitreal injections into mice at postnatal day one. The animals were sacrificed at P26 and the retina fixed in 4%PFA, frozen transverse sections were then immunostained with antibodies against melanopsin, choline acetyl transferase (ChAT), calreinin, calbindin, PKC and Kv4.2. In control retinae, melanosin antibody stained ip-RGC cell bodies and dendrites stratifying in both On and Off layers of the IPL, whereas retinae treated with Mx shows a loss of melanopsin-containing cell bodies and dendrites. In control retinae, ChAT stains starburst amacrine cells with cell bodies in the RGC and INL layers and two distinct bands in the IPL. In Mx treated retinae, most starburst amacrine cells appear to be eliminated along with melanopsin RGCs. Interestingly, if there is a hint of residual melanopsin expressing dendrites remaining, there is also a ChAT expressing cell body and a hint of dendrites in the synaptic layer. In control retinae, calretinin and calbindin antibodies stain a subset of RGCs and amacrine cells and show a characteristic three-layered pattern of dendrites in the IPL. In Mx treated retinae, the calretinin and calbindin layers within the IPL are altered showing an absent or more diffuse labeling pattern in the ON and OFF bands. Antibodies against PKC (staining rod bipolar cells) and Kv4.2 (stains a subset of retinal ganglion cells) do not show an altered staining pattern. These findings suggest that the initial stratification and structural development of synaptic layers in the IPL are altered by Mx treatment.

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Spuz CA, Paolone G, Briscoe S, Bradshaw M, Albin RL, Sarter MF (2011) Deficits in attentional control of balance, mobility, and complex movements in a rat model of early state, multisystem Parkinson disease. Neuroscience 2011 Abstracts 244.02. Society for Neuroscience, Washington, DC.

Summary: In Parkinson disease (PD), basal forebrain cholinergic loss coincides with midbrain dopaminergic neuron loss and contributes to attentional deficits in PD. We hypothesize that these attentional deficits contribute to L-DOPA-insensitive impairments of mobility and postural control in PD. To assess complex movement control, we developed a novel Complex Motor Control Test (CMCT) for rats. The CMCT consists of several 2 m long beams (plank, 13.34 cm width; round rod, 3.81 cm diameter; square rod, 2.54 cm side length), which can be placed at zero, 22.5° or 45° angles in the vertical plane. Rods can rotate at 1 rpm or 10 rpm. A separate ladder apparatus (100 cm long, 7 cm wide, 2 cm between rungs, 5 mm rung diameter) can be placed at zero, 22.5° or 45° angles in the vertical plane and tilted laterally at 15° or 30° angles. Four high-resolution cameras and mirror system record animals’ performances. Rats are habituated by learning that plank traversal allows entry of home compartments containing individual bedding and palatable food. To separately assess attentional performance, we employed our Sustained Attention Task (SAT), including a distractor condition (dSAT). Our initial experiments determined CMCT and SAT performance in three groups: (1) animals with limited (40-60%) loss of cortical cholinergic afferents following immunotoxin 192-IgG saporin basal forebrain lesions (SAP); (2) animals with dopaminergic deafferentation following 6-OHDA dorsal striatal lesions (6-OHDA); (3) animals with both types of deafferentation (DUAL). SAT performance was dramatically impaired in SAP and DUAL animals. Control animals rapidly traversed angled and rotating rods and angled and tilted ladders. Deafferented animals were able to traverse the plank at all angles as effectively as control animals. Cholinergic lesions robustly impaired animals’ ability to maintain balance on the rods, to re-adjust posture on and traverse rotating rods, and had falls (into a net) or dismounts more frequently than control animals. These data reveal unexpectedly striking impairments in complex gait and movement control resulting from loss of corticopetal cholinergic neurons. These results support the hypothesis that basal forebrain cholinergic cell loss in PD contributes to complex posture and movement control deficits.

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Rossi CA, Lehmkuhle MJ, Dudek FE (2011) Evidence that focal interneuron lesions in the hippocampus may lead to a model of epileptogenesis in the mouse. Neuroscience 2011 Abstracts 249.09. Society for Neuroscience, Washington, DC.

Summary: A selective loss of part of the overall population of GABAergic interneurons is a seminal component of many forms of human epilepsy, and is manifest in many animal models of acquired epilepsy, including those based on chemoconvulsant-induced status epilepticus. The current study specifically tests the hypothesis that partial interneuron loss in the dorsal CA1 area of the hippocampus induces epileptiform activity, and the subsequent hypothesis that interictal-like spikes and seizures progressively worsen during the following weeks and months. Focal interneuron lesions were made by intra-hippocampal injection of SSP-Saporin into dorsal CA1 in the hippocampus of GAD67-GFP transgenic mice. Chronic recording electrodes were implanted at the injection site, and local field potentials (LFPs) were monitored continuously during video recording for several weeks. LFP recordings were analyzed for the occurrence of inter-ictal-like paroxysmal events (hippocampal sharp waves of 50-100 msec), and frank seizures. Although interneuron lesions alone were seen to generate inter-ictal-like activity within several days following surgery, full-blown seizure activity was not observed until several weeks later. The current data suggest that disruption of the local GABAergic interneuron population may be a key event that triggers alteration of neural networks in the hippocampus, leading to paroxysmal events and ultimately seizures. The delay in onset suggests other factors besides interneuron loss play a role in the generation of seizures and the development of epilepsy. Thus, loss of local inhibition may be a necessary, but not sufficient condition for epileptogenesis.

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Ljubojevic V, De Rosa E, Luu P (2011) Cholinergic modulation of cross-modal attentional orienting. Neuroscience 2011 Abstracts 294.07. Society for Neuroscience, Washington, DC.

Summary: We modified the classic cued target detection paradigm, using odor cues to predict a visual target, to examine cholinergic modulation of attentional control signals. It has been proposed that without ACh top-down processing will inappropriately dominate in the presence of a low validity cue, i.e., the cue will still drive attention. Thus, we reduced central cholinergic influences in rats after they acquired high validity cues to examine whether top-down processing would dominate even when these same cues changed to a lower validity. The validity effect (VE = invalid cue RT – valid cue RT) is thought to measure the ability to reorient attention, i.e., it reflects the time a subject needs to disengage from an invalidly cued location and shift attention to the actual target location. We trained 8 male Long-Evans rats until they reached the stable performance under baseline conditions: cue validity (CV) = 100%, target duration = 1s. Then we simultaneously manipulated the rats’ cholinergic system and cue validity within a testing session in a 3×3 repeated measures design. The three drug conditions were: muscarinic antagonist scopolamine (0.2mg/kg), muscarinic antagonist methylscopolamine as a peripheral nervous system control (0.2mg/kg), and saline. CV in each session was set to 100%, 75%, or 50%. In sessions with the lower cue validity of 75% and 50%, rats with scopolamine showed the predicted higher validity effect when compared to their performance with the control drugs. Based on the increased VE in scopolamine condition, we conclude that ACh plays a role in attentional orienting when cue and target are presented in a different sensory modality. We hypothesize that scopolamine may have exacerbated the top-down expectations from the cue and increased the validity effect. Thus, we are collecting data from rats that had selective cholinergic lesions of the nucleus basalis magnocellularis, which provides ACh input into the neocortex, with the cholinergic immunotoxin 192 IgG-saporin to support this pharmacological effect. We expect that NBM-ACh-lesioned rats will also have an increased validity effect as the CV decreases relative to the sham-lesioned rats. Also, we will collect the data from 8 additional rats to increase the statistical power of the experiment.

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