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Effects of temperature on intracellular [Ca²⁺] in trout atrial myocytes

Holly A. Shiels^1,*, Matti Vornanen² and Anthony P. Farrell¹

¹ Simon Fraser University, Biological Sciences, Burnaby, British Columbia, V5A 1S6, Canada
² University of Joensuu, Department of Biology, PO Box 111, 80101 Joensuu, Finland

View larger version (12K): [in a new window]   Fig. 1. Action potential (AP) waveforms used to elicit L-type Ca2+ channel current (ICa) and intracellular Ca2+ concentration ([Ca2+]i). The APs were recorded by current-clamp in a separate series of experiments (Shiels et al., 2002a). AP waveforms were obtained by averaging recordings from 6-10 cells at each temperature. The relatively depolarized resting potential is typical for isolated atrial myocytes from rainbow trout (Shiels et al., 2000) and reflects both a low density of the inward rectifier K+ current (IK1) and the lack of cholinergic tone (Vornanen et al., 2002). The inter-pulse holding level during subsequent AP stimulation was -80 mV. The solid line shows the dimensions of the 500 ms square (SQ) pulse.

View larger version (15K): [in a new window]   Fig. 4. This figure is an example of a recording for a single myocyte at 21°C. Part (A) shows intracellular Ca2+ concentration ([Ca2+]i) and part (B) shows the corresponding L-type Ca2+ channel current (ICa) at each stimulus change. Each panel is marked according to the frequency and waveform used to elicit currents and transients. The first five transients at each temperature were elicited by square pulses at 0.2 Hz (SQ0.2Hz). The middle five transients were elicited by SQ pulses applied at a physiologically relevant frequency for the test temperature (SQ1.4Hz), but with a long pulse duration (500ms). Together these factors contribute to the dramatic rise in diastolic Ca2+ levels. The last five transients were elicited by temperature- and frequency-dependent action potentials (AP1.4Hz). The dotted line in the lower panel indicates 0mV. Leakage correction was employed at SQ0.2Hz, but not at SQ1.4Hz or AP1.4Hz.

View larger version (13K): [in a new window]   Fig. 2. The effect of temperature on intracellular Ca2+ concentration ([Ca2+]i) and L-type Ca2+ channel current (ICa) in trout atrial myocytes. (A) The effect of temperature on [Ca2+]i during SQ0.2Hz, SQphysiol and APphysiol (the physiological frequencies were: 0.6 Hz at 7°C, 1.0 Hz at 14°C and 1.4 Hz at 21°C). The increase in the resting level of the transient at SQphysiol reflects the increase in diastolic Ca2+ concentration (see Fig. 4, Table 3). (B) Current recordings of ICa under the same conditions. Peak current amplitude was calculated as the difference between the peak inward current and the current recorded at the end of the depolarizing pulse. The currents elicited at SQ0.2Hz were leakage corrected using the P/N procedure of the software (Clampex, Axon Instruments). Leakage correction was not employed at SQphysiol or APphysiol. Mean data and statistical analysis is given in Tables 1,2,3 and in Fig. 3. SQ0.2Hz, square pulses at a frequency of 0.2 Hz; SQphysiol, square pulses at physiological frequency; APphysiol, action potential at physiological frequency.

View larger version (19K): [in a new window]   Fig. 3. The effect of temperature and stimulus change on (A) the peak current density of L-type Ca2+ channel current (ICa) and (B) the charge density of ICa in trout atrial myocytes. Dissimilar letters indicate significant effects of temperature within each stimulus treatment (analysis of variance, Student—Newman—Keuls test). * indicates a significant decrease in ICa during AP stimulation at all temperatures and with each successive stimulus protocol at 21°C. indicates greater ICa charge density at 7°C during AP stimulation compared with SQ stimulation. Values are means ± S.E.M.; N=7 for 7°C and 21°C, and N=8 for 14°C.