It has long been debated whether changes in Ca2+ are involved in insulin-st
imulated glucose uptake in skeletal muscle. We have now investigated the ef
fect of insulin on the global free myoplasmic Ca2+ concentration and the ne
ar-membrane free Ca2+ concentration ([Ca2+](mem)) in intact, single skeleta
l muscle fibers from mice by using fluorescent Ca2+ indicators. Insulin has
no effect on the global free myoplasmic Ca2+ concentration. However, insul
in increases [Ca2+](mem) by approximate to 70% and the half-maximal increas
e in [Ca2+](mem) occurs at an insulin concentration of 110 microunits per m
i. The increase in [Ca2+](mem) by insulin persists when sarcoplasmic reticu
lum Ca2+ release is inhibited but is lost by perfusing the fiber with a low
Ca2+ medium or by addition of L-type Ca2+ channel inhibitors. Thus, insuli
n appears to stimulate Ca2+ entry into muscle cells via L-type Ca2+ channel
s, Wortmannin, which inhibits insulin-mediated activation of glucose transp
ort in isolated skeletal muscle, also inhibits the insulin-mediated increas
e in [Ca2+](mem). These data demonstrate a new facet of insulin signaling a
nd indicate that insulin-mediated increases in [Ca2+](mem) in skeletal musc
le may underlie important actions of the hormone.