Expression of functional melanocortin-4 receptor in the hypothalamic GT1-1cell line

Mutations in the melanocortin-4 receptor (MC4-R) cause obesity in both mice and humans, and the receptor is presumed to have an important role in the regulation of energy homeostasis. The MC4-R is expressed in discrete sets o f neurons in the central nervous system, and thus it has been technically d ifficult to study the regulation of expression and the signaling mechanisms of this receptor. We report here a neuronal cell line that exhibits endoge nous functional expression for the MC4-R. Initially, RT-PCR analysis showed the presence of MC4-R RNA in the hypothalamic GT1-1 and GT1-7 cells. In ad dition, GT1-7 cells expressed melanocortin-3 receptor while the GT1-1 subcl one specifically expressed predominantly the MC4-R RNA. High-affinity bindi ng sites were demonstrated in the GT1-1 and GT1-7 cells for NDP-alpha melan ocyte-stimulating hormone (MSH; K-i = 1.1 x 10(-10) and 1.8 x 10(-10) M) an d agouti-related protein (AGRP; K-i = 1.548 x 10(-9) and 1.663(-9) M). alph a -MSH-stimulated CAMP production in GT1-1 cells with an EC50 of 2.2 x 10(- 8) M, and CAMP production was inhibited in the presence of AGRP, an endogen ous antagonist of the MC4-R. Stimulation of gonadotropin-releasing hormone (GnRH) secretion was achieved with 1 nM to 1 muM concentrations of NDP-alph a -MSH while no GnRH secretion was observed when the GT1-1 cells were treat ed with AGRP. The data presented here show that GT1-1 cells specifically ex press a functional MC4-R that couples to GnRH release. Copyright (C) 2001 S . Karger AG, Basel.