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J. Biol. Chem., Vol. 266, Issue 3, 1410-1414, Jan, 1991
TJ Shuttleworth and JL Thompson
Several recent studies of intracellular Ca2+ concentration ([Ca2+]i using
fluorescent indicators have involved experiments performed at
nonphysiological temperatures, frequently room temperature. In the studies
reported here, a reduction in temperature was shown to influence the
measurement of [Ca2+]i in two major ways. The first is an effect on the
value for the Kd (apparent dissociation constant) used in the calculation
of [Ca2+]i. The Kd values for indo-1 and fura-2 were found to change with
temperature in a manner analogous to the related compounds
[ethylene-bis(oxyethylenenitrilo)]tetraacetic acid and bis-(o-
aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid. Experiments showed that
the use of previously published Kd values (measured at 37 degrees C) in
experiments at room temperature results in errors of at least 25% in the
calculated [Ca2+]i. The second arises from the differential temperature
sensitivities of the physiological processes that make up any particular
value of [Ca2+]i. While resting [Ca2+]i was relatively unaffected by
reduced temperature (27 degrees C, 13 degrees C), changes in [Ca2+]i
following stimulation were profoundly influenced and in a way that was not
simply predictable. Analysis of the responses indicated that the observed
effects resulted largely from a temperature- dependent reduction in the
rate of Ca2+ removal from the cytosol together with an additional
component, namely a marked reduction in Ca2+ entry, at the lower
temperature (13 degrees C). The net result was observed as changes in the
initial rise in [Ca2+]i on stimulation, together with more profound, and
complex, effects on the sustained elevation in [Ca2+]i following
stimulation. This latter component was markedly increased at 27 degrees C,
but was decreased in cells at 13 degrees C.
Effect of temperature on receptor-activated changes in [Ca2+]i and their determination using fluorescent probes
Department of Physiology, University of Rochester School of Medicine and Dentistry, New York 14642.
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