PHARMACOLOGICAL CHARACTERIZATION OF THE NUCLEOTIDE RECEPTORS THAT MOBILIZE CA2+ IONS IN HUMAN PARATHYROID CELLS

We have used the fluorescent probe fura-2 to perform m agonist studies of the receptor(s) that mobilizes Ca2+ ions in response to extracellular ATP in human parathyroid cells. Extracellular ATP induced Ca2+ responses in both normal and adenomatous parathyroid cells. Activation resulted in an initial small transient response during which Ca2+ ions were released from intracellular stores, followed by a prominent plateau response during which Ca2+ ions entered the cells from the extracellular fluid. The responses exhibited moderate desensitization upon repeated stimulation with ATP, and the ratio of the plateau to the peak response remained constant for any given group of activated cells. The baseline intracellular calcium concentration was 100 +/- 4.3 nM (mean +/- S.E.M., n=3). Following maximal activation by extracellular ATP it rose to a peak of 684 +/- 457 nM (n=3) and a plateau level of415 +/- 9.9 nM (n=3). We examined the effects of a variety of nucleotide species. The order of potency was: adenosine, AMP<alpha,beta-methylene ATP<ADP<ATP approximate to UTP. In the concentration range 1-1000 mu M, UTP (the concentration of agonist inducing a half-maximal response, EC(50)=2.4 mu M) was slightly more potent than ATP (EC(50)=3.6 mu M), and the two nucleotides evoked similar maximal responses. In the concentration range 0.01-1.0 mu M, however, there was a clear difference in the behaviour of the two nucleotides. In particular, ATP, but not UTP, evoked responses that suggested the presence of a second receptor of higher potency but markedly lower efficacy. Responses to 2-methylthioATP (2-MeSATP) in the concentration range 0.01-100 mu M defined a dose-response curve that matched the lower concentration component of the ATP dose-response curve. Together, these data suggest that two receptor species are involved in the Ca2+ responses. One is a nucleotide/P-2U receptor that responds to ATP and UTP, which are approximately equipotent agonists; 2-MeSATP is only a weak agonist with respect to this receptor. The other belongs to the P-2Y receptor subclass. It responds to 2-MeSATP, a selective agonist at P-2Y receptors, and to ATP, but not to the pyrimidine nucleotide UTP.