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The objectives of the present study were to determine the effect of nicotine on MAPK signaling and on the proliferation of AR42J cells as well as to assess the relationship between MAPK activation and exocrine secretion in these cells. AR42J cells were incubated with nicotine and analyzed for the activation of MAPK by Western blot analysis using their respective antibodies and confirmed by immunohistochemistry. The effect of nicotine on cell proliferation was determined by the spec trophotometric method, and cell function was assessed by cholecystokinin (CCK)-stimulated amylase release into the culture medium. Nicotine at a dose of 100 µM induced phospho-ERK½ activation maximally in 3 mm compared with untreated cells. Furthermore, immunofluorescence study confirmed the nicotine-induced increase in translocation of phospho-ERK½ to the nucleus. Activation of phospho-ERK½ was inhibited by an ERK½ pathway inhibitor but not by a nicotine receptor antagonist. At the same dose, there was significantly enhanced proliferation of AR42J cells until 72 h without toxic effect, as the percentage of lactate dehydrogenase release remained unchanged. Other MAPKs, c-Jun NH2-terminal kinase ½ and p38 MAPK, were not affected by nicotine treatment. At a nicotine dose of 100 µM, the CCK-stimulated release of amylase was maximal at 6 mm, and, although a nicotinic receptor antagonist inhibited this response, it was not inhibited by the ERK½ pathway inhibitor. We conclude that nicotine treatment induced activation of ERK ½ and increased the proliferation of AR42J cells. The data further indicate that MAPK signaling by nicotine is independent of the secretory response. [ABSTRACT FROM AUTHOR]

Glucose-induced insulin secretion by pancreatic β-cells depends on membrane depolarization and [Ca2+]i increase. We correlated voltage- and current-clamp recordings, [Ca2+]i measurements, and insulin reverse hemolytic plaque assay to analyze the activity of a thapsigargin-sensitive cationic channel that can be important for membrane depolarization in single rat pancreatic β-cells. We demonstrate the presence of a thapsigargin-sensitive cationic current, which is mainly carried by Na+. Moreover, in basal glucose concentration (5.6 mM), thapsigargin depolarizes the plasma membrane, producing electrical activity and increasing [Ca2+]i. The latter is prevented by nifedipine, indicating that Ca2+ enters the cell through L-type Ca2+ channels, which are activated by membrane depolarization. Thapsigargin also increased insulin secretion by increasing the percentage of cells secreting insulin and amplifying hormone secretion by individual β-cells. Nifedipine blocked the increase completely in 5.6 mM glucose and partially in 15.6 mM glucose. We conclude that thapsigargin potentiates a cationic current that depolarizes the cell membrane. This, in turn, increases Ca2+ entry through L-type Ca2+ channels promoting insulin secretion. [ABSTRACT FROM AUTHOR]