Abstract List for 2018 meeting

#503: Presenilin loop region regulates GSK3β mediated functions on motor proteins during axonal transport

Banerjee, Rupkatha ; Rudloff, Zoe ; Naylor, Crystal ; Gunawardena, Shermali ;

Department of Biological Sciences, SUNY University at Buffalo, College of Arts and Sciences;

Long distance transport within axons is essential for neuronal function and viability, and transport defects have been implicated in Alzheimer’s disease. One possible mechanism by which transport defects could occur is by improper regulation of molecular motors. Previously we found that reduction of Presenilin (PS) stimulated amyloid precursor protein vesicle motility. Reduction of Glycogen synthase kinase 3β (GSK3β) also showed a similar effect. Excess GSK3β caused axonal transport defects and increased motor binding to membranes, while reduction of PS decreased active GSK3β and motor binding to membranes. Further, we identified that functional PS and the catalytic loop region of PS is essential for the rescue of GSK3β-mediated axonal transport defects. Disruption of PS loop (PSΔE9) or expression of the non-functional PS variant, PSD447A, failed to rescue axonal blockages in vivo. Active GSK3β associated with and phosphorylated kinesin-1 in vitro. Current analysis is focused on identifying the GSK3β –mediated phosphorylation sites in kinesin to evaluate their functional significance in controlling motor activity during axonal transport. Together our observations propose a scaffolding mechanism for PS in which the loop region sequesters GSK3β away from motors for the proper regulation of motor function. These findings are important to uncouple the complex regulatory mechanisms that likely exist for motor activity during axonal transport in vivo.

#542: Long non-coding RNAs: junk DNA or diamonds in the rough?

Farkas, Michael H ;

Ross Eye Institute, Department of Ophthalmology, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

The term “junk DNA” was coined in the 1970’s as a reference to regions of the human genome that do not encode mRNA, and therefore is considered non-functional. Since then, multiple classes of non-coding RNAs have been discovered, and, importantly, been shown to have important regulatory functions, and implications in disease pathogenesis. Our lab focuses on a more recently discovered class of non-coding RNAs called long non-coding RNAs (lncRNAs). While largely understudied, lncRNAs can be generally classified as transcriptional regulators. They serve important roles in pluripotency, development, and cellular function. As such, they are becoming widely recognized as important contributors to a wide range of diseases. In the eye, lncRNA expression and functional studies are in their infancy. Our lab has shown that roughly 1,500 lncRNAs are expressed in the retinal pigment epithelium (RPE). We have performed whole transcriptome sub-cellular localization analyses which demonstrate nearly 1,000 lncRNAs enriched in the nucleus and 400 in the cytoplasm, with the remaining showing equal enrichment between the compartments. Using these data, we have identified lncRNAs that bind to epigenetic-regulating proteins, as well as novel lncRNAs that are expressed from loci identified to be considered high risk markers for age-related macular degeneration. Our results suggest lncRNAs have potentially important roles in regenerative medicine, as well as providing novel targets for AMD gene therapy strategies.

#504: Perispinal Targeting of Central TNF Is Antinociceptive in Rats Experiencing Chronic Pain

LaMacchia, Zach M ; Abidi, Asif H ; Ghandili, Mehrnoosh ; Ignatowski, Tracey A ;

Pathology and Anatomical Sciences , SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

Overexpression of tumor necrosis factor-alpha (TNFα) induces hyperalgesia during neuropathic pain (NP) states through its actions of enhancing nociceptive afferent excitability at the site of injury. TNFα also dysregulates neurotransmission in the hippocampus, altering connectivity with corticolimbic regions that process the sensory and emotional components of pain. In this study, we determined whether perispinal (PS) targeting central TNFα would be antinociceptive, and if there was an effect on the perception of pain. NP was induced in male Sprague-Dawley rats using the chronic constriction injury (CCI) model, and thermal hyperalgesia was monitored every other day post-surgery. On day-8 post-surgery, rats were randomized into groups that received a PS injection of an anti-TNF antibody (ab), control IgG isotype ab, or the positive control group that did not receive PS treatment. The affective component, or perception of pain was assessed using conditioned place preference (CPP) to analgesia. Rats experiencing pain were conditioned by being confined in one of three chambers having specific sensory and tactile cues after an injection of saline, and then being confined in a different chamber following injection of amitriptyline (10 mg/kg, IP), a known analgesic. Rats that formed a preference for the amitriptyline-paired chamber have formed a CPP. We found that rats given the PS TNF ab injection experienced alleviation of pain the day following the injection and on the final experiment day (Day 8 vs 9_PreAmi p<0.001; Day 8 vs 10 p<0.001), and did not form a CPP, due to anti-TNF ab-induced alleviation of pain. Rats given the PS IgG ab only had acute alleviation of pain following amitriptyline injection (Day 8 vs 9_PostAmi p<0.001) and did form a CPP (IgG Pre vs Post p<0.03). Our results indicate that the antinociceptive effect of perispinal targeting of central TNF inhibits the perception of pain associated with NP. UB-MDRF (ZL)

#489: Phenotypic and Mechanistic Characterization of a Mouse Model of 16p11.2 Duplication Syndrome

Rein, Benjamin A ;

Department of Physiology and Biophysics, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

Copy number variations (CNVs) at position 11.2 on human chromosome 16 (16p11.2) are associated with a range of debilitating neuropsychiatric outcomes that vary by individual case. Microduplications at 16p11.2 are often coupled with diagnoses of Autism spectrum disorder, intellectual disability, developmental delay and epilepsy, though other phenotypes such as bipolar disorder and schizophrenia have also been reported. A mouse model carrying the 16p11.2 microduplication has been developed (16p11.2^dp/+), though little testing has been done to examine the behavioral phenotype of these animals or examine the neurological underpinnings. As a result, the underlying molecular mechanisms remain poorly understood. The current study thus intended to (a) define the behavioral phenotype of 16p11.2^dp/+mice, (b) investigate the underlying neurobiological mechanisms and attempt to establish links between molecular and behavioral phenotypes, and (c) identify and test potential therapeutic intervention strategies.

#479: Structural correlates of agonist action at nicotinic receptors

Tripathy, Sushree ¹; Zheng, Wenjun ¹; Auerbach, Anthony ²;

¹Department of Physics, ²Department of Physiology and Biophysics, SUNY University at Buffalo;

The amplitude and shape of synaptic responses depend on the energetics of agonist binding and receptor conformational change. Neuromuscular nicotinic acetylcholine receptors (AChRs) are 5 subunit, ligand-gated ion channels that toggle between resting ⇄ active conformations (R⇄R*) that have low⇄high affinity for agonists and a closed⇄open pore. Receptors are allosteric proteins are molecular machines that convert chemical energy from ligand binding into kinetic energy of a global ‘gating’ conformational change. Previously, single-channel electrophysiology experiments showed that for a series of agonists related structurally to the neurotransmitter ACh, binding to R is energetically approximately half of binding to R*. We show that the R/R* binding energy ratio determines the binding-to-gating energy conversion efficiency. Our primary goal is to understand the physical basis of the increase in affinity (chemical binding energy) that is the basis for receptor activation by agonists. We parameterized R and R* binding site structures occupied by a quaternary amine agonist (ACh, TMA, CCh or choline) or the azabicycloamine agonist Epx. After equilibration by using MD, a binding energy was calculated. In R*, the agonist binding pocket is smaller and rotated anti-clockwise, the ligand has a reversed orientation and is closer to the pocket center (distance d_x), and loop C is displaced outward. Of 8 structural parameters that differ between R and R* conformations, d_x was the most-correlated with experimentally-measured binding energy and could account approximately for all agonist actions (affinity, efficacy and efficiency). Indeed, accurate dose-response curves could be calculated from d_x values alone. The contraction and rotation of the binding pocket upon activation resemble at small-scale gating rearrangements of the extracellular domain of related receptors and, hence, could be the trigger for the full gating transition of the receptor.

#534: Cellular and Molecular mechanisms of axon degeneration

Babetto, Elisabetta ¹; Beirowski, Bogdan ²;

¹Department of Pharmacology and Toxicology, ²Department of Biochemistry, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

The Beirowski lab studies mechanisms of axon survival and degeneration. Axons are affected in several neurodegenerative diseases, during aging, and after exposure to toxic chemicals. In most cases, axon loss is responsible for the overt symptoms of these conditions (for example neuropathic pain after chemotherapy treatment). We aim to understand the cell non-autonomous, as well as the cell autonomous mechanisms governing axon survival in health and disease, and how axons respond to injuries. Having published a pioneer study in Nat neuroscience describing the metabolic support offered by Schwann cells to axons, we now expand our investigation to the central nervous system. Here we found that altering metabolic pathways in oligodendrocytes causes a deterioration of axon health during aging in mice, suggesting a role of this central nervous system glia in supporting axons, without affecting myelination. We are exploring which metabolic pathways are involved in this intercellular relationship. Likewise we are also interested in the neuron-autonomous mechanisms of axon degeneration and regeneration, expanding on our initial finding that loss of an E3 ubiquitin ligase abolishes the preconditioning injury response in mice. This block the regenerative reprogramming in the peripheral nervous system, providing insights into the necessity of this pathway for proper axon regeneration after insult.

#511: Modulation of functional neural circuits – stimulation and recording from a cerebral organoid

Bhattacharya, Mahasweta M ¹; Freedman, David ²; Stachowiak, Ewa ³; Stachowiak, Michal ⁴; Dutta, Anirban ⁵;

¹Biomedical Engineering, SUNY University at Buffalo; ²Anatomical Sciences, ³Pathology and Anatomical Sciences, ⁵Biomedical Engineering, SUNY University at Buffalo, School of Medicine and Biomedical Sciences; ⁴Pathology and Anatomical Sciences, Biomedical Engineering;

Electrophysiology can be a powerful technique in deciphering functional microcircuits based on the evoked responses. In this exploratory work, we developed a bidirectional stimulation and recording system using tetrodes and Intan RHD2132 amplifier board for cerebral organoids. We hypothesized that evoked responses generated by activated neurons in the vicinity of the stimulating electrode would reveal neuronal circuit maturity. METHODS Cerebral organoids were generated using a modified protocol published earlier [1]. We stimulated a 48-day old cerebral organoid using a platinum/iridium wire (diameter: 25 µm) and recorded using a polyimide-coated nickel-chrome tetrode wire (diameter: 50µm). We conducted spike detection and computational analysis of the tetrode data using custom code in Matlab. RESULTS Spontaneous neuronal activity was not detected. Computational analysis of the evoked responses (see Figure: boxplot of spikes) revealed correlated tetrode data in the 48-day old cerebral organoid. DISCUSSION We showed that continuous online monitoring of tetrode spike data is feasible to detect neuronal activity however correlated data indicated weak connections and lack of circuit maturation. Furthermore, in the mBCI, spontaneous activity triggered stimulation is possible with our mBCI to guide “Hebbian” learning and microcircuit maturation as future work

#505: Electrochemical characterization of chemogenetically modulated dopamine in the olfactory tubercle

Bhimani, Rohan V ¹; Bass, Caroline E ²; Park, Jinwoo ³;

¹Center for Neuroscience, ²Department of Pharmacology and Toxicology, ³Biotechnical and Clinical Laboratory Sciences, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

The selective targeting of specific neuronal subtypes using chemogenetic techniques, such as Designer Receptors Exclusively Activated by Designer Drugs (DREADDs), has facilitated the understanding of the functional roles of complex brain circuits. While DREADDs are a powerful tool for transient and repeated manipulation of neurons, how activation of excitatory or inhibitory DREADDs affects neurotransmitter dynamics (release and clearance) is poorly understood. In this study, we used a combinatorial viral targeting system to restrict DREADD expression to dopamine (DA) neurons in the ventral tegmental area (VTA)/substantia nigra (SN) of wild-type rats. We then employed in vivo fast-scan cyclic voltammetry (FSCV) to determine how systemic administration of Clozapine-N-oxide (CNO), a biologically inert ligand for DREADDs, modulates DA transmission in the olfactory tubercle (OT), an important limbic structure located in the ventral-most part of the ventral striatum that is implicated in mediating the rewarding effects of drugs. Through immunohistochemical and electrochemical evidence, we demonstrated selective viral targeting of DA neurons and determined that CNO dose-dependently (0.3 - 6.0 mg/kg, i.p) activates DREADDs, leading to excitation and/or inhibition of DA release evoked by electrical stimulation of the VTA/SN in urethane-anesthetized rats. These results will facilitate the understanding of DA neurons in essential brain functions, as well as establish guidelines for the use of DREADDs in behavioral studies.

#515: Role of anterior-hypothalamic vasopressin in play and prosocial USVs in female juvenile rats

Brown, Lauren M ¹; Schatz, Kelcie C ¹; John-Vanderpool, Simone ²; Paul, Matthew J ¹;

¹Department of Psychology, ²The Institute for Strategic Enhancement of Educational Diversity (iSEED), SUNY University at Buffalo, College of Arts and Sciences;

Uncovering the neural circuitry that directs normative social development is crucial for identifying candidate pathways impacted by neurodevelopmental disorders that disrupt social function. The neuropeptide arginine vasopressin (AVP) has been implicated in the regulation of social development, including regulation of juvenile social play and prosocial ultrasonic vocalizations (USVs), however the neural circuitry through which AVP acts is not known. The anterior hypothalamus (AH) has been implicated in social play behavior in Syrian Hamsters – blocking AVP’s action at this site decreases play in juvenile males (Cheng & Delville, 2009). However, the AVP neural circuitry of Syrian hamsters is distinct from other mammals, and it is not known whether AH AVP regulates play in other species or in females. Nor is it known whether AH AVP regulates other prosocial behaviors during development. In the present experiment, we tested whether the stimulatory role of AH AVP in juvenile social play generalizes to female juvenile rats, and to prosocial USVs. Juvenile female Long-Evans rats received infusions of a V1aR antagonist or vehicle into the AH and were tested 2 to 3 h later in a twenty-minute social interaction test with an age-matched, sex-matched, untreated conspecific. Similar to findings in male Syrian hamsters, female rats receiving infusions of the V1aR antagonist exhibited fewer play behaviors than vehicle-treated rats. Antagonist-treated rats also emitted fewer USVs than the vehicle-treated controls. Hence, blocking endogenous AVP in the AH impacts multiple social behaviors in female juvenile rats. These findings support the hypothesis that AH AVP is an important regulator of juvenile social development across species, sexes, and prosocial behaviors.

#536: Sex-specific regulation of ERalpha in the social behavior network of juvenile Siberian hamsters

Carroll , Quinn E ¹; Kalinowski, Leanna ²; Barrett, Abigal ¹; Paul , Matthew J ¹;

¹Department of Psychology, SUNY University at Buffalo, College of Arts and Sciences; ²Research Institute on Addictions, SUNY University at Buffalo;

The prepubertal ovary is often said to be quiescent.  However, our experiments using seasonally-breeding Siberian hamsters have found that photoperiod regulates juvenile social and reward-associated behaviors, in part through a prepubertal ovarian mechanism. Long day lengths (LDs) inhibit, whereas short day lengths (SDs) and prepubertal gonadectomy facilitate, juvenile social play (males and females) and novelty seeking (females only). Photoperiod could regulate ovarian contributions to juvenile behavior by altering the production/secretion of ovarian hormones or by altering the sensitivity of the juvenile brain to these hormones. In this experiment, we tested the hypothesis that photoperiod alters the sensitivity of juvenile female hamsters to estradiol by regulating estrogen receptor-α (ERα) levels in the amygdala, a central node in the social behavior network with connections to the mesolimbic reward pathway. Male and female hamsters reared in a LD or SD were sacrificed at postnatal day 30, and their brains processed by immunohistochemistry for ERα. ERα-immunoreactive (ir) cells in the amygdala were largely restricted to the medial amygdala. LD-reared females had more ERα-ir cells in the posterior medial amygdala than LD-reared males. Notably, this prepubertal sex difference was absent in SD-reared hamsters due to a selective decrease in ERα-ir cells in SD-reared females. No sex or photoperiod differences were detected in the anterior medial amygdala. These findings are consistent with our hypothesis that photoperiod regulates the behavior of female juveniles by altering the sensitivity to prepubertal estradiol. These findings add to a growing literature that challenges the current view of a quiescent prepubertal ovary.