[Ca2+]i-reducing action of cAMP in rat pancreatic β-cells: involvement of thapsigargin-sensitive stores.

In the present study, we examined the ability of adenosine 3',5'-cyclic monophosphate (cAMP) to reduce elevated levels of cytosolic Ca2+ concentration ([Ca2+]i) in pancreatic β-cells. [Ca2+]iand reduced pyridine nucleotide, NAD(P)H, were measured in rat single β-cells by fura 2 and autofluorescence microfluorometry. Sustained [Ca2+]ielevation, induced by high KCl (25 mM) at a basal glucose concentration (2.8 mM), was substantially reduced by cAMP-increasing agents, dibutyryl cAMP (DBcAMP, 5 mM), an adenylyl cyclase activator forskolin (10 μM), and an incretin glucagon-like peptide-1-(7-36) amide (10-9 M), as well as by glucose (16.7 mM). The [Ca2+]i-reducing effects of cAMP were greater at elevated glucose (8.3-16.7 mM) than at basal glucose (2.8 mM). An inhibitor of protein kinase A (PKA), H-89, counteracted [Ca2+]i-reducing effects of cAMP but not those of glucose. Okadaic acid, a phosphatase inhibitor, at 10-100 nM also reduced sustained [Ca2+]ielevation in a concentration-dependent manner. Glucose, but not DBcAMP, increased NAD(P)H in β-cells. [Ca2+]i-reducing effects of cAMP were inhibited by 0.3 μM thapsigargin, an inhibitor of the endoplasmic reticulum (ER) Ca2+ pump. In contrast, [Ca2+]i-reducing effects of cAMP were not altered by ryanodine, an ER Ca2+-release inhibitor, Na+-free conditions, or diazoxide, an ATP-sensitive K+ channel opener. In conclusion, the cAMP-PKA pathway reduces [Ca2+]ielevation by sequestering Ca2+ in thapsigargin-sensitive stores. This process does not involve, but is potentiated by, activation of β-cell metabolism. Together with the known [Ca2+]i-increasing action of cAMP, our results reveal dual regulation of β-cell [Ca2+]iby the cAMP-signaling pathway and by a physiological incretin.