1 NAADP has been shown to be a potent calcium-releasing second messenger in a wide variety of cell types to date. However, research has been hampered by a lack of pharmacological agents, with which to investigate NAADP-induced calcium release, and by the molecular identity of its cellular target protein being unknown. 2 In the present paper, the sea urchin egg model was used to investigate whether triazine dyes, which can act as nucleotide mimetics, can bind to the NAADP receptor, induce Ca 2 þ release and be used for affinity chromatography of the receptor. 3 Indeed, all the triazine dyes tested (Reactive Red 120 (RR120), Reactive Green 19 (RG19), Reactive Green 5 (RG5), Cibacron Blue 3GA and Reactive Yellow 86) displayed micromolar affinities, except for Reactive Orange 14. Furthermore, unlike NAADP, RR120, RG19 and RG5 did not bind in an irreversible manner. 4 The compound that displayed the highest affinity, RR120, was tested in a 45 Ca 2 þ efflux assay. This compound released Ca 2 þ via the NAADP receptor, as shown by the ability of subthreshold NAADP concentrations to inhibit this release. Furthermore, heparin and ruthenium red were unable to block RR120-induced Ca 2 þ release. 5 We have also shown that RG5 and RG19, immobilised on resins, retain the ability to bind to the receptor, and that this interaction can be disrupted by high salt concentrations. As a proof of principle, we have shown that this can be used to partially purify the NAADP receptor by at least 75-fold. 6 In conclusion, triazine dyes interact with the NAADP receptor, and this could be exploited in future to create a new generation of pharmacological tools to investigate this messenger and, in combination with other techniques, to purify the receptor.

Evidence is given that demonstrates the reliability of the bihyperbolic equation, proposed by Plá-Delfina and Moreno, in fitting the correlation between absorption rate constants (ka) found in the small intestine and in the colon of the living anesthetized rat, and partition constants (1/RF-1), for a series of phenylalkylamines, a group of compounds which differ largely from others which have been tested. Emphasis is laid on the nonexistence of an optimum of lipophilicity for intestinal absorption/partition correlation: This feature makes inapplicable the probabilistic approaches to the reported data.

The uptake of zinc as acexamic acid salt in the small intestine of the anaesthetized rat was shown to be a two-phase process in normal animals. The first phase is rapid mucosal binding which satisfies the Freundlich isotherm equation and which involves about 30 per cent of the initially perfused zinc. The second phase was characterized as an apparent absorption step which obeys Michaelis-Menten and first-order combined kinetics, with the following parameters: Vm = 6.51 mg h-1; Km = 2.96 mg; ka = 0.306 h-1. In largely non-saturated conditions, an apparent global rate constant of about 2.50 h-1 was calculated. No significant interference due to endogenous zinc excretion into the small intestine was observed during the absorption period. In zinc-deficient animals, the two phases were not so well characterized. Binding was non-linear and apparent absorption efficiency was much greater at high zinc concentrations, so no evidence of saturable kinetics was found, thus confirming the hypothesis of a homeostatic zinc regulation mechanism.