Diadenosine polyphosphates in insulin-secreting cells: interaction with specific receptors and degradation.

Abstract

A role of diadenosine polyphosphates as second messengers was suggested for insulin-secreting cells. It has not yet been investigated whether specific receptors for these compounds exist and how these extracellular compounds and their degradation products may contribute to insulin release. Specific saturable binding sites for diadenosine polyphosphates exist in INS-1 cells and rat pancreatic islets. In INS-1 cells, the rank order of diadenosine polyphosphates displacing [3H]Ap4A from binding sites was Ap4A = Ap5A >Ap3A = Ap6A. Binding was specific, since suramin was not able to displace the binding; adenosine, ATP, UTP, alpha,beta-methylene ATP, beta,gamma-methylene ATP, ADP-betaS, 2-methylthio ATP, and pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) were able to displace [3H]Ap4A from its binding sites. Insulin release was investigated in INS-1 cells. Perifusion experiments showed an increase in insulin release stimulated by the diadenosine polyphosphates in the presence of 8.3 mmol/l glucose; in static incubations (90 min), however, insulin release was inhibited dose dependently by the four diadenosine polyphosphates. This discrepancy might be due to the instability of the compounds. [3H]Ap4A was degraded in the extracellular medium to mainly adenosine and low concentrations of ATP, ADP, AMP, and inosine (half-maximal degradation after 25 min). The insulin stimulatory effect is due to the original compounds (acute perifusion experiments), and the insulin inhibitory effect (static incubation experiments) is due to the production of inhibitory compounds, such as adenosine, in the medium. Small amounts of intact [3H]Ap4A, but mainly [3H]ATP, accumulated in the cells within 20 min. The uptake of labeled compounds is dependent on an intact metabolism and intact receptor internalization. This data indicates that 1) specific bindings sites for diadenosine polyphosphates exist in INS-1 cells and rat pancreatic islets mediating insulin release; 2) the receptors involved in INS-1 cells may be diadenosine polyphosphate receptors, albeit others, such as P2X-receptors, cannot be ruled out; and 3) diadenosine polyphosphates, and mainly their degradation products in the extracellular space, are to a high degree accumulated within cells with unknown function. Thus, diadenosine polyphosphates are worth being investigated more closely in physiological and pathophysiological terms.