(50) DESENSITIZATION OF MITOCHONDRIAL Ca2+ AND INSULIN SECRETION RESPONSES IN THE B-CELL

Pierre Maechler, Eleanor D. Kennedy, Haiyan Wang and Claes B. Wollheim
Division de Biochimie Clinique, Department of Internal Medicine, University Medical Center, CH-1211 Geneva 4, Switzerland

The role of mitochondria in the desensitization of insulin secretion was investigated. In rat pancreatic bð-cells, both insulin secretion and mitochondrial [Ca2+] ([Ca2+]m) rises were desensitized following two challenges with the mitochondrial substrate methyl-succinate. In the bð-cell line INS-1 similar results were observed when a 5min interval separated two 5min pulses. In contrast, ATP generation monitored in luciferase-expressing INS-1 cells was stimulated to the same extent during both exposures to methyl-succinate. Succinate activates the electron transport chain at complex II. As a consequence, the mitochondrial membrane hyperpolarizes, promoting ATP synthesis and Ca2+ influx into the mitochondria through the uniporter. The mitochondrial desensitization was further studied in permeabilized INS-1 cells. Increasing extramitochondrial [Ca2+] from 100nM to 500nM enhanced succinate oxidation 4 fold. At 500nM Ca2+, 1mM succinate caused a blunted [Ca2+]m rise upon the second, compared to the first, stimulation. Succinate hyperpolarized the mitochondrial membrane both during first and second applications. This suggests that the uniporter itself, rather than the respiratory chain, is desensitized. Accordingly, mitochondrial stimulation was then performed by only increasing the free [Ca2+] in the intracellular buffer. As a consequence, the second exposure to 1.3µM Ca2+ showed a complete desensitization of the transient increase in [Ca2+]m above the equilibration value between the two compartments, i.e., the cytosol and the mitochondrial matrix. These results emphasize the key role of the mitochondria not only in the stimulation of insulin secretion, but also in its desensitization.