Presentations and Posters of Interest Society for Neuroscience Meeting (2015) 34.01/A100. Estradiol rapidly attenuates ORL-1 receptor-mediated inhibition of proopiomelanocortin neurons via Gq-coupled, membrane-initiated signaling *K. M. CONDE1, C. MEZA2, M. KELLY3, K. SINCHAK4, E. WAGNER2; 1Grad. Col. of Biomed. Sci., 2Col. of Osteo. Med. of the Pacific, Western Univ. of Hlth. Sci., Pomona, CA; 3 Dept. of Physiol. & Pharmacol., Oregon Hlth. and Sci. Univ., Portland, OR; 4California State University, Long Beach, Long Beach, CA Ovarian estrogens act through multiple receptor signaling mechanisms that converge on hypothalamic arcuate nucleus (ARH) proopiomelanocortin (POMC) neurons. A subpopulation of these neurons project to the medial preoptic nucleus (MPN) to regulate lordosis. Orphanin FQ/nociception (OFQ/N) via its opioid-like receptor (ORL-1) regulates lordosis through direct actions on these MPN-projecting POMC neurons. Based o an ever-burgeoning precedence for fast steroid actions, we explored whether estradiol excites ARH POMC neurons by rapidly attenuating inhibitory ORL-1 signaling in these cells. Experiments were carried out in hypothalamic slices prepared from ovariectomized female rats injected one-week prior with the retrograde tracer Fluorogold into the MPN. During electrophysiologic recordings, cells were held at or near -60 mV. Post-hoc identification of neuronal phenotype was determined via immunohistofluorescence. In vehicle-treated slices OFQ/N caused a robust outward current/hyperpolarization via activation of GIRK channels. This OFQ/N-induced outward current was attenuated by 17-β estradiol (E2, 100nM). The 17α enantiomer of E2 had n effect. The OFQ/N-induced response was also attenuated by an equimolar concentration of E2 conjugated to BSA. In addition, the ability of E to diminish OFQ/N responsiveness was blocked by the co-administration of the estrogen receptor (ER) antagonist ICI 182,780 (1μM). The attenuating effect of E2 was mimicked by the membrane ER (mER) ligand STX (10nM) and the ERα agonist PPT (1μM), but not the GPR30 agonist G1 (3μM) or the ERβ agonist DPN (3μM). Moreover, the phospholipase C (PLC) inhibitor U73122 (20μM) restored the OFQ/N-induced outward current in the presence of E2, whereas the inactive analog U73343 (20μM) was without effect. Finally, the protein kinase C (PKC) inhibitor NPC 15437 (30μM) abrogated the estrogenic impairment of the OFQ/N-induced outward current, whereas the PKC activator PDBu (1μM) per se attenuated the OFQ/N response. These collective actions were observed in a substantial number of MPN-projecting ARH neurons positive for various markers of POMC neurons. The results reveal an ORL-1 receptor mediated inhibition of POMC neurons that is negatively modulated by estradiol. The estrogenic attenuation is stereoselective; membrane delimited, mediated via the Gq- coupled mER and ERα activation, and involves signaling through PLC and PKC. This disinhibition of POMC neurons is critical for the subsequent expression of sexual behavior in the female. ____________________________________________________________________________________ 1 Page 34.04/A103. Synapse-specific persistent activation of VTA kappa opioid receptors following acute stress *A. M. POLTER1, R. CHEN1, R. M. ST. LAURENT2, J. A. KAUER1; 1Mol. Pharmacology, Physiology, and Biotech., 2Neurosci., Brown Univ., Providence, RI Stressful experiences drive many adaptive and maladaptive behaviors, and even acute stressors can have lasting behavioral consequences. Emerging evidence shows that dopaminergic neurons in the ventral tegmental area (VTA) are an important locus in stress. We previously identified a long-term potentiation of GABAergic synapses onto these neurons (LTPGABA) that is blocked by acute stress (Graziane et al, Neuron, 2013). Administration of a kappa opioid receptor (KOR) antagonist (norBNI) in vivo prevents the block of LTPGABA. Intra-VTA injection of the KOR antagonist also prevents reinstatement of cocaine seeking by acute stress, suggesting that KOR-mediated regulation of VTA inhibitory plasticity may play a role in stress-induced drug seeking. Our recent work shows that a single five minute cold water swim stress blocks LTPGABA for at least five days. Surprisingly, blocking KORs with norBNI even well after stress restores LTPGABA, and cocaine self-administration is prevented even when norBNI is administered after stress (Polter et al, Biological Psychiatry, 2014). Here we show that the long-lasting block of LTPGABA by stress is due to changes in the KOR itself. While bath application of an inverse agonist (norBNI, 10 nM) rescues LTPGABA in slices from stressed animals, bath application of a neutral antagonist (6--naltrexol, 10 µM) does not (LTP magnitude: norBNI after stress=144± 18% of baseline, 6--naltrexol after stress=99±8% of baseline; p<0.05). These results suggest that LTPGABA is blocked by constitutive activation of KORs rather than by persistently elevated dynorphin, which would be blocked by both drugs. Transient activation of KORs was sufficient to induce a lasting blockade of LTPGABA, as a KOR agonist (U50488, 5 mg/kg) blocked LTPGABA for 5 days (LTP: saline=140±10% of baseline, 1 day post U50488=108±5% of baseline, 5 days post U50488=99±9% of baseline). The activation of KORs by stress is synapse specific, as bath application of norBNI did not potentiate excitatory synapses on either dopaminergic (IPSC amplitude after norBNI: control=92±6% of baseline, FSS=94±2% of baseline) or GABAergic VTA neurons (IPSC amplitude after norBNI: control=109±4% of baseline, FSS=112±3% of baseline). Our results show that a single exposure to acute stress or KOR activation both cause long-lasting changes in activity of KORs specifically at GABAergic synapses in the VTA. ____________________________________________________________________________________ 34.05/A104. Variable sensitivity to morphine mediated Ferritin Heavy Chain upregulation in cortical neuronal subpopulations *B. S. NASH1, K. TARN2, J. H. PITCHER2, O. MEUCCI2; 2 2Pharmacol. and Physiol., 1Drexel Univ. Col. of Med., Philadelphia, PA Page Current HIV therapies have dramatically improved patients’ disease progression and quality of life, but HIV-associated neurocognitive disorder remains prevalent and challenging problem to address in the clinic. Many HIV+ patients also abuse drugs, with opioid use being particularly common as sharing needles represents an important avenue of infection. These patients can show accelerated progression of cognitive impairment, the mechanism of which is currently under investigation. Our lab has shown that morphine and other mu-opioids upregulate Ferritin Heavy Chain (FHC) in CNS neurons, which is associated with reduction of downstream signals of the homeostatic chemokine receptor CXCR4 via its natural ligand CXCL12. This results in a host of adverse effects including reduced dendritic spine density and is correlated with enhanced cognitive decline in humans and animal models of HAND. Our experiments aim to gain insights into this novel mechanism of mu-opioid mediated CXCR4 regulation, and thereby potential future targets for novel HAND therapies. Morphine upregulates FHC in cortical neurons in a mu-opioid/g-protein dependent manner, while astrocytes seem to be unaffected in vitro even though they express the mu-opioid receptor. This suggests exclusivity for particular CNS cell types in their ability to upregulate FHC via opioids. FHC upregulation specifically occurs in the cytoplasm of neuronal cells as demonstrated by fractionation and confocal imaging studies, which allows it to potentially interact with CXCR4 on the cell membrane. Preliminary imaging studies show that certain neurons are more susceptible to FHC upregulation than others, and that GABA transporter-1 expressing neurons represent one of these susceptible populations. Current work is focused o exploring FHC expression after morphine treatment in inhibitory and excitatory neuronal subpopulations both in vitro and in vivo. Pilot studies suggest variability amongst neuronal subpopulations in that calretenin expressing interneurons do not upregulate FHC after morphine, but interestingly have higher basal levels of FHC compared to calretenin negative cortical neurons. Future studies will characterize additional neuronal populations. These experiments suggest that mu-opioid usage may cause specific deficits in inhibitory neuronal circuits via FHC upregulation and subsequent CXCR4 blockade, which may induce or sustain particular features of HAND such as excitotoxicity, or other neurochemical adaptations leading to cognitive impairment. ___________________________________________________________________________________ 34.14/B5. Localization of AGRP immunoreactivity in the mouse hippocampus that expresses eGFP- tagged GHSR1a B. GONZALEZ, *M. ISOKAWA; Dept. of Biomedicine, Univ. Texas-Brownsville, Brownsville, TX The ghrelin receptor (GHSR1a) is expressed highly in the hippocampus (Mani et al., 2014) and contributes to learning and memory (Diano et al., 2006). However, in spite of intense investigation with immunohistochemical techniques and transgenic eGFP expression, what types of hippocampal neurons express GHSR1a is not well understood. Available evidence suggests hippocampal principal neurons (glutamatergic pyramidal cells and dentate granule cells) are likely the target of ghrelin (Diano et al., 2006; Cuellar and Isokawa, 2011). However, in the hypothalamus, GHSR1a is specifically localized in 3 AgRP/NPY/GABA-containing neurons (Wang