Calpain activity was measured in the various subfractions of rat myoca
rdia after global ischemia for 60 min or after ischemia followed by 30
min of reperfusion after the chromatographic separation of mu- and m-
calpains. The activity of m-calpain after ischemia and that of mu-calp
ain after reperfusion were both higher than that in the control. The a
ctivity of the endogenous calpain inhibitor calpastatin in 10,000Xg su
pernatant was decreased after both ischemia and ischemia-reperfusion.
The increase in m- and mu-calpain activities was suppressed by pre-isc
hemic perfusion with a synthetic calpain inhibitor, transepoxysuccinyl
-L-leucylamido (4-guanidino) butane (E64d, 100 mu g/ml). After reperfu
sion, the calpain activity in the 10,000Xg pellet was also increased,
which was inhibited by pre-ischemic perfusion with E64d or dimethylsul
foxide (a solvent for E64d) or by reperfusion with 1 mmol/L ethylenegl
ycol bis (beta-aminoethylether)-N, N, N', N'-tetraacetic acid. SDS-pol
yacrylamide gel electrophoresis revealed the proteolysis of several pr
oteins, including fodrin, in the 10,000Xg and 100,000Xg pellet fractio
ns after ischemia and reperfusion, some of which were confirmed to be
in vitro substrates of calpain. The creatine kinase release during the
reperfusion was also partially inhibited by E64d or dimethylsulfoxide
. Thus, calpain activity in the soluble or particulate fractions was a
ltered during ischemia or reperfusion, and appeared to be implicated i
n the proteolysis of the membrane proteins, which may contribute to my
ocardial