Proceedings of The Physiological Society

University of Bristol (2005) J Physiol 567P, PC59

Poster Communications

Electrophysiological evidence for functional melanocortin 3 and 4 receptors in mouse hypothalamic arcuate neurones.

Smith, Mark; Withers, Dominic; Barsh, Gregory; Ashford, Michael;

1. Pathology and Neuroscience, University of Dundee, Dundee, United Kingdom. 2. Centre for Diabetes and Endocrinology, University College London, London, United Kingdom. 3. Genetics and Pediatrics, Stanford University, Stanford, CA, USA.


Alpha-melanocyte stimulating hormone (α-MSH) is made and released from a population of hypothalamic arcuate neurones that express pro-opiomelanocortin (POMC). α-MSH binds to melanocortin (MC) 3 and 4 receptors and reduces body weight and food intake. Conversely, a different arcuate neuronal population make and release agouti-related peptide (AGRP), an inverse agonist to MC 3/4 receptors. Recently we identified a novel arcuate neuronal population that regulates energy homeostasis, by the expression of green fluorescent protein (GFP) induced by the rat insulin 2 promoter (RipCre) transgene (Choudhury et al. 2005). Here we demonstrate that RipCre neurones lie within the hypothalamic melanocortin neuronal network. Hypothalamic arcuate neurones were identified in acute brain slices by epiflorescence and differential interference contrast microscopy. Mice (8-16 weeks) were humanely killed. Whole cell current-clamp recordings were made from GFP positive neurones driven by POMC or RipCre promoters, as previously described (Choudhury et al., 2005). All data are expressed as means ± S.E.M and statistical significance determined by paired t-test.. RipCre and POMC neurones spontaneously fired sodium-mediated action potentials at resting membrane potentials (-50 ± 1 mV, n = 62 & -50 ± 1 mV, n = 50 respectively). A mixed MC3/4 receptor agonist (MTII, 100 nM) depolarized RipCre (-53 ± 3 to -48 ± 2 mV, P < 0.01; n = 9) and POMC (-53 ± 3 to 49 ± 3 mV, P < 0.01; n = 6) neurones and this was associated with an increase in action potential firing. In contrast, AGRP (10 nM) reduced spike firing frequency in RipCre and POMC neurones by cellular hyperpolarization (RipCre: -47 ± 4 to -63 ± 5 mV, P < 0.04, n = 4; POMC: -46 ± 2 to -48 ± 1 mV, P < 0.01, n = 7). The selective MC3 (γ-Trp-MSH, 10 nM) and MC4 (Cyclo (β-Ala-His-D-Phe-Arg-Trp-Glu)-NH2; 10 nM) receptor agonists depolarized POMC (MC3: -44 ± 2 to -40 ± 3 mV, P < 0.03, n = 4; MC4: -49 ± 1 to -47 ± 1 mV, P < 0.04, n = 4) and RipCre (MC3: -54 ± 5 to -45 ± 5 mV, P < 0.04, n = 5; MC4: -54 ± 3 to -51 ± 3 mV, P < 0.07, n = 6) neurones. Note that RipCre neuron depolarization by the MC4 receptor agonist was not statistically significant at the 95% level, although firing frequency increased from 3.2 ± 0.7 to 4.0 ± 0.5 Hz (P < 0.04). These data suggest that although expression of MC4 receptors is low in the arcuate nucleus (Liu et al., 2003), both MC 3 and 4 receptor subtypes are functional in these neurones.

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