Effects of na+k+2cl cotransport inhibition on myocardial na and ca
during ischemia and reperfusion.
Anderson, S. E., C. Z. Dickinson, H. Liu, and P. M. Cala.
Department of Human Physiology, University of California, Davis,
California 95616-8644
APStracts 2:0319C, 1995.
In the context of the "pump-leak" hypothesis (37), changes in
myocardial intracellular Na (Nai) during ischemia and reperfusion
have historically been interpreted to be the result of changes in Na
efflux via the Na/K pump. We investigated the alternative hypothesis
that changes in Nai during ischemia are the result of changes in the
Na "leak" rather than changes in the pump. More specifically, we
hypothesize that the increase in Nai during ischemia is in part the
result of increased Na uptake mediated by Na/H exchange. Further, we
present data consistent with the interpretation that the Na+K+2Cl
cotransporter is active (or alternatively, displaced from
equilibrium) during ischemia and may contribute an additional Na
efflux pathway during reperfusion. Thus inhibition of Na efflux via
Na+K+2Cl cotransport during ischemia and reperfusion could result in
increased Nai and therefore decreased force driving Ca efflux via
Na/Ca exchange and ultimately increased intracellular Ca
concentration ([Ca]i). Nai (meq/kg dry wt) and [Ca]i (nM) were
measured in isolated Langendorff-perfused rabbit hearts using NMR
spectroscopy. Except during 65 min of ischemia, hearts were perfused
with HEPES buffered Krebs-Henseleit solution equilibrated with 100%
O2 at 23 C and pH 7.4+/-0.05 . During ischemia Nai rose from 16.6+/
-0.3 to 62.9+/-5.1 (_Nai nearly equal to 46) meq/kg dry wt and
decreased during subsequent reperfusion (mean+/-SEM, n=3). In order
to measure Na uptake ("leak") in the absence of efflux via
the Na/K pump, in all of the protocols described below the perfusate
was nominally K-free solution containing 1mM ouabain for 10 min prior
to ischemia and during 30 min of reperfusion. After K-free perfusion
Nai rose from 20.2+/-0.5 to 79.1+/-5.3 (_Nai nearly equal to 59)
meq/kg dry wt (n=3) during ischemia and decreased during K-free
reperfusion. When amiloride (1mM) was added to the K-free perfusate
to inhibit Na/H exchange, Nai rose from 16.3+/-0.9 to 44.7+/-5.1
(_Nai nearly equal to 28) meq/kg dry wt (n=3) during ischemia, i.e.
amiloride decreased Na uptake. When bumetanide (20 [mu]M) was added
to the nominally K-free perfusate to inhibit Na+K+2Cl cotransport,
Nai rose from 22.5+/-3.9 to 83.8+/-13.9 (_Nai nearly equal to 61)
meq/kg dry wt (n=3) during ischemia and did not decrease during
reperfusion, i.e. bumetanide inhibited Na recovery during reperfusion
(p<0.05 compared to bumetanide-free). For the same protocol, the
presence of bumetanide resulted in increased [Ca]i during ischemia
and reperfusion (p<0.05); these increases in [Ca]i are
interpreted to be the result of increased Nai (see above). Thus the
results are consistent with the hypotheses.
Received 6 September 1994; accepted in final form 25 August 1995.
APS Manuscript Number C529-4.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 23 September 1995.