Increased Intracellular Calcium Is Required for Spreading of Rat Islet β-Cells on Extracellular Matrix

Increased Intracellular Calcium Is Required for Spreading of Rat Islet β-Cells on Extracellular Matrix Domenico Bosco 1 , Carmen Gonelle-Gispert 1 , Claes B. Wollheim 2 , Philippe A. Halban 1 and Dominique G. Rouiller 1 1 Research Laboratories Louis-Jeantet and the 2 Division of Clinical Biochemistry, University of Geneva, Geneva, Switzerland Abstract Rat islet β-cells spread in response to glucose when attached on the matrix produced by a rat bladder carcinoma cell line (804G). Furthermore, in a mixed population of cells, it has been observed previously that spread cells secrete more insulin acutely in response to glucose, compared with cells that remain rounded. These results suggest bi-directional signaling between the islet β-cell and the extracellular matrix. In the present study, the role of increased intracellular free Ca 2+ concentration [Ca 2+ ] i as an intracellular step linking glucose stimulation and β-cell spreading (inside-out signaling) was investigated. Purified rat β-cells were attached to this matrix and incubated under various conditions known to affect [Ca 2+ ] i . The effect of glucose on β-cell spreading was mimicked by 25 mmol/l KCl (which induces calcium influx) and inhibited by diazoxide (which impairs depolarization and calcium entry) and by the l -type Ca 2+ channel blocker SR-7037. When a 24-h incubation at 16.7 glucose was followed by 24 h at 2.8 mmol/l, β-cells that had first spread regained a round phenotype. In the presence of thapsigargin, spreading progressed throughout the experiment, suggesting that capture of calcium by the endoplasmic reticulum is involved in the reversibility of spreading previously induced by glucose. Spreading was still observed in degranulated β-cells and in botulinum neurotoxin E–expressing β-cells when exocytosis was prevented. In summary, the results indicate that increased [Ca 2+ ] i is required for the glucose-induced spreading of β-cells on 804G matrix and that it is not a consequence of exocytotic processes that follow elevation of [Ca 2+ ] i . Footnotes Address correspondence and reprint requests to Domenico Bosco, Laboratoires de Recherche Louis-Jeantet, Centre Médical Universitaire, 1, rue Michel-Servet, 1211 Geneva 4, Switzerland. E-mail: domenico.bosco{at}medecine.unige.ch . Received for publication 16 August 2000 and accepted in revised form 31 January 2001. BSA, bovine serum albumin; [Ca 2+ ] i , intracellular free Ca 2+ concentration; DMEM, Dulbecco’s minimum essential medium; ECM, e