References

Blumenthal SA, Maness EBL, Fadel JR, Burk JA (2019) Effects of an orexin-2 receptor agonist on attention in rats following loss of cortical cholinergic projections. Neuroscience 2019 Abstracts 418.06. Society for Neuroscience, Chicago, IL.

Summary: Deterioration to the basal forebrain cholinergic system (BFCS) is linked to age-related cognitive impairment, specifically to the pathology of Alzheimer’s disease (AD). Animals with BFCS damage perform poorly on learning, memory, and attention tasks, indicating cognitive deficits. The orexin neuropeptide system, comprised of two neuropeptides (orexin A and orexin B), has also been implicated in the cognitive decline associated with aging, likely due to the role of orexins in promoting attention. Two orexin receptor subtypes exist, orexin 1 (Ox1R) and orexin 2 (Ox2R). Studies have examined the effects of stimulation and blockage of both receptors together and Ox1R alone on attention; but no studies have examined the role of Ox2Rs in attention through the use of Ox2R agonists. Ox2Rs may be implicated in attentional processes and the loss of orexin neurons seen in age-related cognitive decline. In order to examine the role of Ox2Rs in attention following BFCS deterioration, the present study administered the Ox2R agonist, YNT-185, to rats given intrabasalis infusions of either saline (n = 12) or 192 IgG saporin (n=11), an immunotoxin which selectively destroys the BFCS. Animals received infusions of YNT-185 to the lateral ventricle (LV) in doses of 0, 1, 10, and 100nM across four separate sessions and performance was then assessed on a sustained attention task requiring discrimination between signal and non-signal trials through lever presses. The 100nM dose of YNT-185 improved attentional performance, as compared to the 0nM dose, for rats given the immunotoxin, but worsened performance for rats given saline lesions. YNT-185 may be efficacious in aiding attentional function in animals with vulnerable cholinergic systems but may lead to overexcitation for those with intact cholinergic function.

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Clement MK, Pimentel CS, Swaine JA, Pimentel AJ, Hutchins D, McGaughy JA (2019) Dissociable attentional effects of dopaminergic and cholinergic lesions to the anterior cingulate cortex. Neuroscience 2019 Abstracts 418.11. Society for Neuroscience, Chicago, IL.

Summary: Prior work from our lab has shown that excitotoxic lesions to the anterior cingulate cortex (ACC) impairs the ability of rats to filter certain types of distracting stimuli (Newman and McGaughy 2011). Specifically, rats with lesions of the ACC cannot filter distractors that have been made salient through pairing with reinforcement. In contrast, these same subjects can filter distracting stimuli that have not been predictive of reward. The present study investigates the effects of neuromodulator specific lesions of the same region to determine how specific neuromodulators contribute to the attentional function of ACC. Cholinergic or dopaminergic deafferentation of the ACC was achieved using either 192 IgG saporin (n=10) or dopamine transporter saporin (n=10). Lesions were restricted to the rostral portion of the area and did not spread to nearby prefrontal sub-regions e.g prelimbic cortex. After lesioning, subjects were tested in an attentional set-shifting task (Birrell and Brown 2000). While both cholinergic and dopaminergic lesions increased distractibility, these deficits were not as severe as those produced after excitotoxic lesions (n= 8). In contrast to excitotoxic lesions, both cholinergic and dopaminergic lesions also impeded formation of an attentional set. Because dopaminergic lesions produced impairments in many stages of the tasks, we hypothesized that these subjects had a more general impairment in stimulus processing. In order to address these broader processing impairments, we analyzed the data to determine whether lesioned rats showed more sensitivity to novel stimuli, or made more perseverative errors. The implications of these data for understanding the unique contributions of acetylcholine and dopamine to attentional processing in the ACC will be discussed.

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Bhyrapuneni G, Benade V, Daripelli S, Kamuju V, Shinde A, Abraham R, Nirogi R, Jasti V (2019) SUVN-G3031, histamine H3 receptor inverse agonist preclinical evaluation for the treatment of excessive daytime sleepiness in narcolepsy. Neuroscience 2019 Abstracts 502.07. Society for Neuroscience, Chicago, IL.

Summary: Numerous studies have demonstrated that brain histamine plays a crucial role in maintenance of wakefulness, attention, learning and other cognitive processes. SUVN-G3031, a potent histamine H3 receptor inverse agonist is being developed for the treatment of narcolepsy and other sleep related disorders. SUVN-G3031 is one of the lead molecules with hKi of 8.7 nM and has more than 100 fold selectivity against the related GPCRs. SUVN-G3031 exhibited desired pharmacokinetic properties and brain penetration. SUVN-G3031 blocked R-α-methylhistamine induced water intake and increased tele-methylhistamine levels in brain and cerebrospinal fluid. In the present study, SUVN-G3031 was evaluated in brain microdialysis and rodent models of electroencephalography (EEG). SUVN-G3031 was evaluated in brain microdialysis for evaluation of neurotransmitters like acetylcholine, histamine, dopamine and norepinephrine in male Wistar rats. EEG was used to evaluate the effects on sleep/ wake profile in rats and mice.A single oral administration of SUVN-G3031 produced significant increase in acetylcholine, histamine, dopamine and norepinephrine levels in the cortex. SUVN-G3031 produced no change in the dopamine levels of striatum and nucleus accumbens indicating that SUVN-G3031 may not have addiction liabilities. Narcoleptic-like sleep behavior was observed in rats injected with hypocretin-2-saporin in lateral hypothalamus. SUVN-G3031 produced significant increase in wakefulness with concomitant decrease in rapid eye movement (REM) sleep in these animals. These results are in agreement with EEG studies carried out in healthy male Wistar rats. Results from current studies provide strong evidence for the potential of SUVN-G3031 in the treatment of excessive daytime sleepiness associated with narcolepsy. First in human, Phase 1 studies for SUVN-G3031 are completed under US IND and SUVN-G3031 has shown desirable pharmacokinetic profile with safety and tolerability in healthy human volunteers. Phase 2 study for the treatment of excessive daytime sleepiness associated with narcolepsy is currently ongoing in USA.

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Walker JR, Ong A, Detloff MR (2019) Role of nociceptive afferent input on forelimb reaching and grasping behaviors in the spinal cord injured rat. Neuroscience 2019 Abstracts 572.09. Society for Neuroscience, Chicago, IL.

Summary: Individuals with spinal cord injury (SCI) suffer a loss of motor and sensory function. The current standard of care to recover fine motor control is rehabilitation focused on a combination of range of motion, aerobic, and strength training (ST). However, limited research has been conducted to determine the role of nociceptive afferent inputs from muscle on spinal plasticity and/or recovery of function. Using a rodent model of SCI strength training rehabilitation, we determined that motor training not only improves forelimb strength and fine motor function but also can modulate the development of neuropathic pain, suggesting that improvements in reaching and grasping may be due, in part, to plasticity of nociceptive afferents. To further explore this, Sprague-Dawley rats received injections of rIB4-conjugated saporin, mu p75-conjugated saporin or unconjugated (vehicle) into the cervical dorsal root ganglia unilaterally to eliminate non-peptidergic and peptidergic nociceptors. There is an uninjured cohort and a group with unilateral C5 SCI. Von Frey and Hargreaves’ tests were performed at baseline and several time points post-injection to assess the effcacy of the nociceptive elimination. Several measures of forelimb strength were recorded over time including the isometric pull task, a single pellet retrieval task and the Montoya staircase test. To confirm the depletion of peptidergic and non-peptidergic nociceptors following saporin injection and/or SCI, cervical DRGs and spinal cords were stained with antibodies against CGRP and isolectin-B4. An understanding of the role of nociceptors in spinal plasticity and functional motor and sensory recovery of SCI patients will guide future research and refine rehabilitation strategies to further improve their quality of life.

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Harris RB (2019) Leptin receptor activity in the nucleus of the solitary tract increases forebrain leptin sensitivity. Neuroscience 2019 Abstracts 591.04. Society for Neuroscience, Chicago, IL.

Summary: We previously reported that fourth ventricle infusions of leptin that cause weight loss are associated with an increase in hypothalamic phosphorylation of signal transducer and activator of transcription 3 (pSTAT3), a marker of leptin receptor (ObRb) activation, implying an integrated response to central leptin. This study tested the impact of ObRb activity in the nucleus of the solitary tract (NTS) on sensitivity to leptin in the forebrain. Leptin-Saporin (Lep-Sap) injections were used to delete ObR- expressing neurons in the NTS of 300g male Sprague Dawley rats. Controls were injected with Blank-Saporin (Blk-Sap). Loss of NTS ObR was confirmed with RNAScope in situ hybridization and pSTAT3 response to peripheral leptin in representative Lep- Sap rats. Experimental rats were fitted with 3rd ventricle (3V) guide cannula 12 days after Lep-Sap or Blk-Sap injections. Nine days later cannula placement was tested with Angiotensin II and rats were adapted to calorimeter cages for 4 days. Lep-Sap had no effect on body weight. To test leptin responsiveness rats were food deprived for 5 hours and at 5 p.m. they received 3V injections of 0, 0.05, 0.1, 0.25 or 0.5 μg leptin. Food was returned at 6 p.m., the start of the dark period. Each rat received the injections in random order at 4 day intervals. At the end of the experiment NTS pSTAT3 was used to confirm effcacy of Lep-Sap injections. Seven Lep-Sap and 6 control Blk-Sap rats completed the experiment. There was a dose-dependent inhibition of food intake in Blk-Sap rats, but only 0.5 μg leptin inhibited intake of Lep-Sap rats. Intake was inhibited during the 24 hours following injection and was not compensated for so that cumulative intake was inhibited for 60 hours post-injection. Energy expenditure was not different between groups and respiratory exchange ratio tended to follow food intake. These data suggest that leptin- induced inhibition of food intake is mediated by an integrated network involving both the forebrain and hindbrain and that activation of NTS ObRb lowers the threshold for leptin responsiveness in the forebrain.

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Suarez AN, Liu CM, Cortella AM, Noble EN, Kanoski SE (2019) Medial septum cholinergic signaling regulates gastrointestinal-derived vagus sensory nerve communication to the hippocampus. Neuroscience 2019 Abstracts 601.19. Society for Neuroscience, Chicago, IL.

Summary: The vagus nerve delivers bi-directional communication between feeding-relevant gastrointestinal (GI) signals and the brain. Vagal sensory-mediated GI satiation signals, including gastric distension and intra-gastric nutrient infusion, activate neurons in the hippocampus (HPC). Recent work from our lab revealed that selective GI-derived vagal sensory signaling is required for HPC-dependent episodic and visuospatial memory, effects accompanied by reduced dorsal HPC (dHPC) expression of neurotrophic and neurogenic markers. To investigate the neural pathways mediating gut regulation of hippocampal-dependent memory, here we investigate the hypothesis that GI-derived signals communicate to dHPC neurons via cholinergic input from the medial septum, a memory-promoting pathway that is vulnerable to disruption in various degenerative dementia diseases. To explore this putative gut-to-brain pathway, we administered 192IgG-saporin, a neurotoxin that selectively kills cholinergic neurons via apoptosis, in the medial septum to determine whether septal cholinergic neurons regulate vagally-mediated neuronal activation in dHPC. Results revealed that elimination of cholinergic neurons in the MS reduced peripherally-administered cholecystokinin (CCK)-induced c-Fos expression in the dHPC, suggesting that cholinergic inputs from the MS transmit GI-derived signaling to the dHPC. Consistent with this interpretation, dHPC protein expression of vesicular acetylcholine transporter (VAChT), which promotes memory function and acetylcholine release without disrupting other co- released molecules, was significantly reduced in rats with GI-specific vagal sensory ablation via nodose ganglion injections of CCK conjugated to saporin. Collectively these results suggest that GI-derived vagal sensory signaling infuences memory function via enhancement of MS cholinergic signaling to the dPHC.

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Cunningham C, Lowry JP (2019) In vivo monitoring of cholinergic neurotransmission with a microelectrochemical choline biosensor. Neuroscience 2019 Abstracts 614.03. Society for Neuroscience, Chicago, IL.

Summary: Acetylcholine acts as a key neuromodulator within the central nervous system, capable of altering neuronal excitability and coordinating neuronal firing patterns. Conversely, cholinergic neurotransmission plays a crucial role in a variety of cognitive functions, including the encoding of new memories. Cholinergic neuronal loss, and the resulting drop in cholinergic neurotransmission (collectively referred to as hypocholinergia), is closely associated with cognitive dysfunction in a number of chronic neurodegenerative disorders including Alzheimer’s disease. However, conventional analytical techniques for monitoring in vivo cholinergic neurotransmission lack the spatiotemporal resolution required to accurately detect endogenous cholinergic dynamics. Here we validate in mice a Pt-based electrochemical biosensor for selective monitoring of choline, a verified marker of cholinergic transmission. Enzymatic choline biosensors (modified with choline oxidase) were sterotaxically implanted in the medial prefrontal cortex (mPFC) and contralateral dorsal hippocampus (dHPC) of female C57Bl6J mice. Real-time choline current recordings over a period of several days revealed circadian fluctuations in both regions, with extracellular choline levels highest during light phases. Administration of pharmacological compounds known to induce central acetylcholine release, scopolamine (1mg/kg) and amphetamine (4mg/kg), evoked a robust increase in choline current. In contrast, peripheral injection of the reversible acetylcholinesterase inhibitor, donepezil (3mg/kg), produced a marked decrease in recorded choline current. The induction of systemic infammation with bacterial lipopolysaccharide (LPS; 500µg/kg) produced characteristic ‘sickness behaviour’ in mice and evoked a tonic rise in central choline levels in both the mPFC and dHPC. Furthermore, the induction of hypocholinergia in selected mice was preformed via intracerebroventricular injections of murine-p75-saporin immunotoxin (1.2µg). Evoked cholinergic neurotransmission was dramatically attenuated in lesioned (hypocholinergic) mice. Collectively, the data suggests that microelectrochemical choline biosensors may serve as a powerful tool for monitoring cholinergic neurotransmission across a number of behavioural and disease states.

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Lee J, Seo Y, Shin J, Kong C, Na Y, Chang W, Chang J (2019) Increased transplantation efficacy of mesenchymal stem cell by focused ultrasound and improvement of the spatial memory in the 192 IgG-saporin rat model. Neuroscience 2019 Abstracts 048.01. Society for Neuroscience, Chicago, IL.

Summary: Introduction: Stem cell therapy has been found to have therapeutic effects in neurodegenerative disease, but traditional transplant methods, such as parenchymal or intravenous injection, possess limitations like secondary injuries, infection, and low survival rate of stem cells in the brain. Meanwhile, recently the focused ultrasound(FUS) was found to have promising results regarding transplantation of stem cells into the rat brain. However, the mechanism of stem cell transplantation with FUS and possibility of cognitive recovery remain elusive. Therefore, this study investigates a possibility for non-invasive focused ultrasound use in stem cell transplantation into the brain of dementia rat model. Materials & methods: We divided rats into five groups: Normal, Lesion, Cell only, FUS + Cell, and FUS only. We used 192 IgG-saporin for degeneration of basal forebrain cholinergic neuron and it was injected into all rats except for the normal group. After a week, 5p mesenchymal stem cells (MSC: 3*106/200ul) were injected in the tail vein of all rats of the cell only and FUS + Cell group, and the FUS + Cell group received the FUS three hours before cell transplantation. FUS was applied with parameters of 0.25Mpa, 300s (Targeted hippocampal region: AP -3.5, ML ±2). And last, FUS only group was received only FUS without any treatment. Five weeks after transplantation, rats performed the Morris water maze test. Results: MSC were detected in both cell only and FUS + Cell group of the hippocampus region. After comparing FUS+MSC & cell only rats, it was confirmed that FUS increases MSC homing in the sonicated rat’s brain. In addition, the most effective memory restoration occurred in the FUS + Cell group. Moreover, the FUS + Cell group exhibited better recall of the platform position than the other groups. And FUS only group did not recover. Conclusion: Noninvasive FUS can increase the efficacy of stem cell delivery. And memory impairment due to cholinergic denervation can be effectively improved by cell transplantation with FUS. The results of this study suggest possibility of stem cell homing and therapeutic effects of the FUS in dementia rat model. However, further study regarding the function of stem cells transplanted in the brain and a more detailed mechanism of stem cell homing by FUS is needed.

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Grayson VS, Han Y, Guedea AL, Jovasevic V, Gao C, Apkarian A, Radulovic JM (2019) The role of subcortical hippocampal inputs in contextual memory formation. Neuroscience 2019 Abstracts 786.03. Society for Neuroscience, Chicago, IL.

Summary: The role of cortical efferents to the hippocampus in the formation of episodic-like memory is well established, however, less is known about the contribution of subcortical memory circuits to memory. In the present study, we studied the roles of several subcortical inputs into the dorsal hippocampus in mouse models of contextual fear conditioning, extinction, and reinstatement. Fear conditioning was induced by a single exposure of mice to a context followed by foot shock. Subsequently, mice were exposed to daily extinction trials. After significant reduction of freezing, indicating successful extinction, mice were exposed to a brief reminder shock and re-tested in the conditioning context. Circuit manipulations were performed by chemogenetic silencing with the inhibitory designer receptor exclusively activated by designer drugs (DREADD) hM4(Gi) or targeted cholinergic depletion induced by 192 IgG-saporin, at different stages of fear conditioning, extinction, and reinstatement. We identified projection- and neurotransmitter-specific roles of discrete circuits, indicating complex regulation of fear-inducing memories by subcortical afferents.

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Wang S, Kim M, Ong K, Pae E-K, Chung M-K (2019) Nociceptors expressing TRPV1 and trigeminal nucleus neurons expressing NK1 mediate orthodontic pain. Neuroscience 2019 Abstracts 052.10. Society for Neuroscience, Chicago, IL.

Summary: Orthodontic force produces mechanical irritation and inflammation in periodontium, which inevitably accompanies pain. Despite its high prevalence, treatment of orthodontic pain is not effective. Determining detailed neural mechanisms involving peripheral and central nervous system should be critical to improve the management of orthodontic pain. Periodontal ligament is projected by peptidergic nociceptors, which is enriched with transient receptor potential vanilloid 1 (TRPV1), a receptor for capsaicin. Trigeminal subnucleus caudalis (Vc), is critical for relaying orofacial nociceptive signal into brain. A group of second- order neurons in the superficial dorsal horn of Vc express neurokinin 1 receptor (NK1), a receptor for substance P, and receive inputs from peptidergic nociceptors. However, the contribution of these nociceptive neurons to orthodontic pain has not been determined. Orthodontic force of 10g produced reliable tooth movement in mice. Orthodontic pain was evaluated by measuring mouse grimace scale (MGS) and bite force (BF), which could represent spontaneous pain and chewing-evoked pain, respectively. Orthodontic force increased MGS and decreased BF, which peaked at 1d and returned near to sham level at 7d. Using targeted chemical ablation of specific subsets of neurons, we determined the contribution of TRPV1+ nociceptors and NK1+ Vc neurons to orthodontic pain behaviors in mice. Ablation of TRPV1+ nociceptors by injecting resiniferatoxin into trigeminal ganglia significantly attenuated orthodontic force assessed by MGS and BF. Chemical ablation of NK1+ Vc neurons by injecting saporin conjugated with substance P into Vc also significantly reduced the extent of changes in MGS and BF by orthodontic force. These results suggest that TRPV1+ trigeminal nociceptors and NK1+ Vc neurons constitute a major neural pathway for transmission of orthodontic pain, which is a fundamental neural mechanism of orthodontic pain transmission. The new mouse model of orthodontic pain will be useful for mechanistic study to develop novel approaches for painless orthodontics.

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