Neurons synthesizing melanin-concentrating hormone identified by selective reporter gene expression after transfection in vitro: transmitter responses.
Neurons from the lateral hypothalamus that synthesize melanin-concentrating hormone (MCH) play an important role in the regulation of energy homeostasis. Relatively little is known of the cellular physiology and transmitter responses of these neurons, in part because of the difficulty in identifying live MCH cells. Here we use a novel approach of transfection of specific gene constructs with the MCH promoter driving green fluorescent protein (GFP) or red fluorescent protein (dsRed2) in CNS cultures to identify live rat MCH neurons; all neurons expressing the reporter gene showed MCH immunoreactivity, indicating selective expression. MCH neurons had a resting membrane potential of -57.5 +/- 0.6 mV, a linear current-voltage relation and a mean input resistance of 1,013 MOmega. Long depolarizing pulses revealed significant spike frequency adaptation. Functional glutamate and GABA receptors were expressed by MCH neurons. MCH neurons were hyperpolarized by norepinephrine in the presence or absence of tetrodotoxin, suggesting direct inhibition. Orexigenic peptides neuropeptide Y (NPY) and MCH showed no direct effect on membrane potential, input resistance, action potential width, or afterhyperpolarization potential, but inhibited voltage-dependent calcium channels, indicating that MCH neurons expressed both MCH and NPY receptors.
Department of Neurosurgery, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Xiao-Bing
XB
Gao
Prabhat K
PK
Ghosh
Anthony N
AN
van den Pol
D015763:2-Amino-5-phosphonovalerate / Q000494:pharmacology; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione / Q000494:pharmacology; D000818:Animals; D001640:Bicuculline / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D015870:Gene Expression / Q000502:physiology*; D017930:Genes, Reporter / Q000235:genetics*; D007028:Hypothalamic Hormones / Q000096:biosynthesis* / Q000235:genetics*; D007150:Immunohistochemistry; D008543:Melanins / Q000096:biosynthesis* / Q000235:genetics*; D008564:Membrane Potentials / Q000502:physiology; D009474:Neurons / Q000378:metabolism*; D009478:Neuropeptide Y / Q000494:pharmacology; D018377:Neurotransmitter Agents / Q000378:metabolism*; D010641:Phenotype; D010907:Pituitary Hormones / Q000096:biosynthesis* / Q000235:genetics*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D018691:Excitatory Amino Acid Antagonists; D007028:Hypothalamic Hormones; D008543:Melanins; D009478:Neuropeptide Y; D018377:Neurotransmitter Agents; D010907:Pituitary Hormones; C019463:melanin-concentrating hormone; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione; D015763:2-Amino-5-phosphonovalerate; D001640:Bicuculline