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Pharmacological properties of the Ca 2+ ‐release mechanism sensitive to NAADP in the sea urchin egg
Author(s) -
Genazzani A A,
Mezna M,
Dickey D M,
Michelangeli F,
Walseth T F,
Galione A
Publication year - 1997
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1038/sj.bjp.0701295
Subject(s) - ryanodine receptor , sea urchin , inositol trisphosphate , cyclic adp ribose , biochemistry , receptor , chemistry , biophysics , intracellular , voltage dependent calcium channel , biology , inositol , calcium , microbiology and biotechnology , cd38 , stem cell , cd34 , organic chemistry
The sea urchin egg homogenate is an ideal model to characterize Ca 2+ ‐release mechanisms because of its reliability and high signal‐to‐noise‐ratio. Apart from the InsP 3 ‐ and ryanodine‐sensitive Ca 2+ ‐release mechanisms, it has been recently demonstrated that this model is responsive to a third independent mechanism, that has the pyridine nucleotide, nicotinic acid adenine dinucleotide phosphate (NAADP), as an endogenous agonist. The sea urchin egg homogenate was used to characterize the pharmacological and biochemical characteristics of the novel Ca 2+ ‐releasing agent, NAADP, compared to inositol trisphosphate (InsP 3 ) and cyclic ADP ribose (cyclic ADPR), an endogenous activator of ryanodine receptors. NAADP‐induced Ca 2+ ‐release was blocked by L‐type Ca 2+ ‐channel blockers and by Bay K 8644, while InsP 3 ‐ and cyclic ADPR‐induced Ca 2+ ‐release were insensitive to these agents. L‐type Ca 2+ ‐channel blockers did not displace [ 32 P]‐NAADP binding, suggesting that their binding site was different. Moreover, stopped‐flow kinetic studies revealed that these agents blocked NAADP in a all‐or‐none fashion. Similarly, a number of K + ‐channel antagonists blocked NAADP‐induced Ca 2+ ‐release selectively over InsP 3 ‐ and cyclic ADPR‐induced Ca 2+ ‐release. Radioligand studies showed that these agents were not competitive antagonists. As has been shown for InsP 3 and ryanodine receptors, NAADP receptors were sensitive to calmodulin antagonists, suggesting that this protein could be a common regulatory feature of intracellular Ca 2+ ‐release mechanisms. The presence of K + was not essential for NAADP‐induced Ca 2+ ‐release, since substitution of K + with other monovalent cations in the experimental media did not significantly alter Ca 2+ release by NAADP. On the contrary, cyclic ADPR and InsP 3 ‐sensitive mechanisms were affected profoundly, although to a different extent depending on the monovalent cation which substituted for K + . Similarly, modifications of the pH in the experimental media from 7.2 to 6.7 or 8.0 only slightly affected NAADP‐induced Ca 2+ ‐release. While the alkaline condition permitted InsP 3 and cyclic ADPR‐induced Ca 2+ ‐release, the acidic condition completely hampered both Ca 2+ ‐release mechanisms. The present results characterize pharmacologically and biochemically the novel Ca 2+ ‐release mechanism sensitive to NAADP. Such characterization will help future research aimed at understanding the role of NAADP in mammalian systems.