Expression of functional P2-purinergic receptors in primary cultures of human colorectal carcinoma cells

Primary cell cultures of human colorectal carcinomas were established and characterized immunocytochemically. In the isolated cancer cells intracellular Ca2+ concentrations ([Ca2+](i)) were measured by the fura-2 method. Stimulation with either extracellular ATP or UTP caused a biphasic rise of [Ca2+]i in a dose-dependent manner and cross-desensitization between both nucleotides was observed. The rank order of potency was ATP greater than or equal to UTP > ATP-gamma-S > ADP > adenosine which is characteristic for a P-2U-receptor subtype. Selective agonists of P-1-, or P-2X-purinoceptors had no effect on [Ca2+](i). The initial rise in [Ca2+](i) was independent of extracellular calcium [Ca2+](e), whereas the second phase was not observed under [Ca2+](e)-free conditions suggesting a capacitative Ca2+-entry-mechanism. Intracellular Ca2+ mobilization was proven by use of the Ca2+-ATPase inhibitor thapsigargin. P-2U-specific mRNA could be detected by RT-PCR in both colorectal tumor tissues and in the human colorectal cancer cell line HT 29. In HT 29 cells, the hydrolysis-resistant ATP analog ATP-gamma-S inhibited cell proliferation and, also, induced apoptosis in a dose-dependent manner. Thus, human colorectal cancer cells express functional P-2U-receptors which may play a role in the regulation of cell. proliferation and apoptosis. (C) 1998 Academic Press.