Temperature dependence of cloned mammalian and salmonid cardiac Na(+)/Ca(2+) exchanger isoforms

The cardiacNa⁺/Ca²⁺ exchanger (NCX), an importantregulator of cytosolic Ca²⁺ concentration in contractionand relaxation, has been shown in trout heart sarcolemmal vesicles tohave high activity at 7°C relative to its mammalian isoform. Thisunique property is likely due to differences in protein structure. Inthis study, outward NCX currents (I_NCX) of thewild-type trout (NCX-TR1.0) and canine (NCX 1.1) exchangers expressedin oocytes were measured to explore the potential contributions ofregulatory vs. transport mechanisms to this observation. cRNA wastranscribed in vitro from both wild-type cDNA and was injected intoXenopus oocytes. I_NCX of NCX-TR1.0 and NCX1.1 were measured after 3-4 days over a temperature range of 7-30°C using the giant excised patch technique. TheI_NCX for both isoforms exhibitedNa⁺-dependent inactivation and Ca²⁺-dependentpositive regulation. The I_NCX of NCX1.1exhibited typical mammalian temperature sensitivities withQ₁₀ values of 2.4 and 2.6 for peak and steady-statecurrents, respectively. However, the I_NCX ofNCX-TR1.0 was relatively temperature insensitive with Q₁₀values of 1.2 and 1.1 for peak and steady-state currents, respectively.I_NCX current decay was fit with a singleexponential, and the resultant rate constant of inactivation () wasdetermined as a function of temperature. As expected,  decreasedmonotonically with temperature for both isoforms. Although  wassignificantly greater in NCX1.1 compared with NCX-TR1.0 at alltemperatures, the effect of temperature on  was not differentbetween the two isoforms. These data suggest that thedisparities in I_NCX temperature dependencebetween these two exchanger isoforms are unlikely due to differences intheir inactivation kinetics. In addition, similar differences intemperature dependence were observed in both isoforms after-chymotrypsin treatment that renders the exchanger in a deregulatedstate. These data suggest that the differences in I_NCX temperature dependence between the twoisoforms are not due to potential disparities in either theI_NCX regulatory mechanisms or structuraldifferences in the cytoplasmic loop but are likely predicated ondifferences within the transmembrane segments.