ELSEVIER Molecular Brain Research 36 (1996) 127-136 MOLECULAR BRAIN RESEARCH Research report Structure of a parathyroid hormone/parathyroid hormone-related peptide receptor of the human cerebellum and functional expression in human neuroblastoma SK-N-MC cells M. Eggenberger ~, B. Fliihmann a, R. Muff a, M. Lauber h, W. Lichtensteiger b, W. Hunziker c, J.A. Fischer a, W. Born ~'" a Research Laboratory for Calcium Metabolism, Departments of Orthopedic Surgery and Medicine. Unicersity of Zurich, Forch~'tras~e 340, 8008 Zurich, Switzerland h Institute of Pharmacology, Unicersity of Zurich-lrchel, 8057 Zurich, Switzerland c F. Hoffmann-La Roche, Ltd., 4002 Basel, Switzerland Accepted 20 September 1995 Abstract Cloning and functional expression of a cDNA from the human cerebellum revealed a parathyroid hormone/parathyroid hormone-re- lated peptide (PTH/PTHrP) receptor protein of 593 amino acids, identical in sequence to the PTH/PTHrP receptor of the human kidney and an osteoblast-like cell line (Schipani et al., Endocrinology, 132 (1993) 2157-2165). Expression of m RNA hybridizing with the cloned cDNA, indistinguishable in size on Northern blots from a 2.3 kb transcript in kidney and liver, was detected in eight brain areas. In situ hybridization histochemistry in rat brain tissue sections revealed predominant signals in the Purkinje cell layer of the cerebellum and in the mesencephalic nucleus of the trigeminal nerve. In human neuroblastoma (SK-N-MC) cells, stably transfected with the cloned eDNA, hPTH(I-84) and hPTH(1-34) displaced binding of 125 pM [t251][Tyr36}chPTHrP(1-361 to the PTH/PTHrP receptor with IC5, values of 4.0 + 0.6 nM and 2.00 + 0.08 nM, and stimulated cyclic AMP accumulation with ECso values of 0.19 + 0.06 nM and 0.09 + 0.01 nM. respectively. 16 out of 48 cells responded to 100 nM hPTH(I-341 with a 2-10-fold transient increase of cytosolic free calcium concentrations. In conclusion, a PTH/PTHrP receptor, identified in the human cerebellum, has the primary structure of the corresponding receptors of kidney and bone. Expression in human neuroblastoma SK-N-MC cells revealed functional properties indistinguishable from those of non-neuronal tissues. The widespread distribution of PTHrP and its receptor in brain implies biological functions remaining to be elucidated. Keywords." Parathyroid hormone/parathyroid hormone-related peptide receptor; Human brain; Cerebellum; SK-N-MC cell line: Nervous system 1. Introduction Parathyroid hormone (PTH), a polypeptide of 84 amino acid residues, is predominantly produced by the parathy- roid glands. The secretion of PTH is inversely related to extracellular calcium levels. Predominant target organs are the kidney and bone. Evidence for the synthesis and local action of PTH has been reported in the central nervous system [8]. Immunoreactive PTH, and expression of PTH mRNA has been detected in the rat hypothalamus by Northern blot analysis, in situ hybridization and poly- merase chain reaction (PCR) amplification of cDNA re- • Corresponding author. Fax: 1411 (l) 386 16 52. 0169-328X/9~/$15.00 ~" 1996 Elsevier Science B.V. All rights reserved SSDI (~1 f~q-328X(~.~51I)0253 -7 verse transcribed from hypothalamic mRNA [5,211]. Local action of PTH identified in the hypothalamus was sug- gested, based on the observed stimulation of dopamine turnover by intracerebroventricularly (i.c.v.) administered PTH, and perifusion and incubation of hypothalamus tissue slices with micromolar concentrations of PTH [9,10]. PTH stimulated calcium transport into brain synaptosomes of the rat and raised cytosolic calcium concentrations [4,26]. Along these lines, PTH inhibited calcium currents in mouse neuroblastoma cells and the firing rate of neurons in the ventromedial nucleus of the hypothalamus, and i.c.v, ad- ministered PTH prevented urethane-induced hypocalcemia [18,21]. Parathyroid hormone-related peptide (PTHrP), associ- ated with the syndrome of humoral hypercalcemia of