fg-9041 and norbinaltorphimine

Hypothalamic proopiomelanocortin (POMC) neurons and their peptide products mediate important aspects of energy balance, analgesia, and reward. In addition to peptide products, there is evidence that POMC neurons can also express the amino acid transmitters GABA and glutamate, suggesting these neurons may acutely inhibit or activate downstream neurons. However, the release of amino acid transmitters from POMC neurons has not been thoroughly investigated in an intact system. In the present study, the light-activated cation channel channelrhodopsin-2 (ChR2) was used to selectively evoke transmitter release from POMC neurons. Whole-cell electrophysiologic recordings were made in brain slices taken from POMC-Cre transgenic mice that had been injected with a viral vector containing a floxed ChR2 sequence. Brief pulses of blue light depolarized POMC-ChR2 neurons and induced the release of GABA and glutamate onto unidentified neurons within the arcuate nucleus, as well as onto other POMC neurons. To determine whether the release of GABA and glutamate from POMC terminals can be readily modulated, opioid and GABA(B) receptor agonists were applied. Agonists for μ- and κ-, but not δ-opioid receptors inhibited transmitter release from POMC neurons, as did the GABA(B) receptor agonist baclofen. This regulation indicates that opioids and GABA released from POMC neurons may act at presynaptic receptors on POMC terminals in an autoregulatory manner to limit continued transmission. The results show that, in addition to the relatively slow and long-lasting actions of peptides, POMC neurons can rapidly affect the activity of downstream neurons via GABA and glutamate release.

Topics: Adrenocorticotropic Hormone; Analgesics, Opioid; Animals; Channelrhodopsins; Excitatory Postsynaptic Potentials; Female; gamma-Aminobutyric Acid; Glutamic Acid; Hypothalamus; In Vitro Techniques; Inhibitory Postsynaptic Potentials; Light; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Naltrexone; Narcotic Antagonists; Nerve Net; Neurons; Neurotransmitter Agents; Patch-Clamp Techniques; Peptides; Presynaptic Terminals; Pro-Opiomelanocortin; Quinoxalines

Hypothalamic proopiomelanocortin (POMC) neurons release the endogenous opioid beta-endorphin and POMC neuron activity is inhibited by opioids, leading to the proposal that beta-endorphin acts to provide feedback inhibition. However, both intrinsic properties and synaptic inputs contribute to the regulation of POMC neurons such that attributing an autoregulatory role to opioids must include consideration of opioid receptor localization and sensitivity at both presynaptic and postsynaptic sites. In the present study, whole-cell recordings were made in POMC cells in mouse brain slices and the presynaptic and postsynaptic regulation of POMC neurons was examined using selective agonists for mu, kappa, and delta opioid receptors. Activation of mu, but not kappa or delta, receptors induced a direct postsynaptic outward current. Agonists for each of the receptors inhibited the frequency of spontaneous IPSCs. Mu and kappa, but not delta, agonists reduced the amplitude of evoked IPSCs and appeared to colocalize in a significant portion of GABAergic terminals onto POMC neurons. The presynaptic inhibition caused by the mu agonist DAMGO had an EC(50) of 80 nM, whereas the EC(50) was 350 nM when measuring the postsynaptic outward current. This differential sensitivity adds an unexpected component of opioid-dependent feedback regulation, where low levels of opioid receptor activation would likely disinhibit POMC neuron activity and higher concentrations would result in an overall inhibition. The results may help explain why it has been difficult to clearly discern the role that opioids play in the regulation of food intake and other processes involving POMC neurons.

Topics: Analgesics; Analgesics, Opioid; Analysis of Variance; Animals; Benzeneacetamides; Dizocilpine Maleate; Dose-Response Relationship, Drug; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Gene Expression; Hypothalamus; In Vitro Techniques; Inhibitory Postsynaptic Potentials; Luminescent Proteins; Membrane Potentials; Mice; Mice, Inbred C57BL; Mice, Transgenic; Naltrexone; Narcotic Antagonists; Neural Inhibition; Neurons; Patch-Clamp Techniques; Peptide Fragments; Pro-Opiomelanocortin; Pyrrolidines; Quinoxalines; Receptors, Opioid; Somatostatin; Synapses