Identification of MELla melatonin receptors using a peptide specific polyclonal antibody

Abstract

Melatonin plays an important role in regulating arcadian rhythms in various species including man. Its biologic action is mediated through specific receptors (MELR) which have been found in the CNS as well as in other tissues, such as kidney. To gain insight into how melatonin regulates kidney function we sought to identify its renal subcellular localization. Accordingly we prepared a polyclonal antibody to a highly conserved peptide derived from the predicted intracellular loop of the human brain MELla receptor (Neuron 13, 1995). Immunoblotting studies were conducted on brush border (BBM) and basolateral membranes (BLM) from guinea pig kidney as well as human embryonic (HEK) and opossum (OK) kidney cells, and compared with preimmune serum. A specific peptide blockable band of MW 37-38 kDa was detected in BBM, BLM and HEK cells, but not in OK cells. No band was observed with preimmune serum. The presence of this 37-38 kDa band correlated with the presence of high affinity MELR binding sites determined by radioligand binding using iodinated melatonin. These data demonstrate the specificity of the anti MELR antibody. The renal localization of MELR suggests that melatonin may have an affect on cortical tubular function. Further studies are planned to map MELR in other regions of the kidney as well as other epithelia.