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

• Published in: American Journal of Physiology: Cell Physiology Vol. 281; no. 3; pp. C993 - C1000
• Main Authors: Elias, C L, Xue, X H, Marshall, C R, Omelchenko, A, Hryshko, L V, Tibbits, G F
• Format: Journal Article
• Language: English
• Published: United States 01.09.2001
• Subjects: Animals; Calcium - physiology; Cell Membrane - physiology; Chymotrypsin; Cloning, Molecular; DNA, Complementary; Dogs; Female; Heart - physiology; In Vitro Techniques; Mammals; Membrane Potentials - drug effects; Membrane Potentials - physiology; Oocytes - physiology; Patch-Clamp Techniques; Protein Isoforms - genetics; Protein Isoforms - physiology; Recombinant Proteins - metabolism; RNA, Complementary; Sodium - pharmacology; Sodium-Calcium Exchanger - genetics; Sodium-Calcium Exchanger - physiology; Species Specificity; Temperature; Thermodynamics; Trout; Xenopus laevis
• ISSN: 0363-6143
• DOI: 10.1152/ajpcell.2001.281.3.c993

The cardiac Na(+)/Ca(2+) exchanger (NCX), an important regulator of cytosolic Ca(2+) concentration in contraction and relaxation, has been shown in trout heart sarcolemmal vesicles to have high activity at 7 degrees C relative to its mammalian isoform. This unique property is likely due to differences in protein structure. In this study, outward NCX currents (I(NCX)) of the wild-type trout (NCX-TR1.0) and canine (NCX 1.1) exchangers expressed in oocytes were measured to explore the potential contributions of regulatory vs. transport mechanisms to this observation. cRNA was transcribed in vitro from both wild-type cDNA and was injected into Xenopus oocytes. I(NCX) of NCX-TR1.0 and NCX1.1 were measured after 3-4 days over a temperature range of 7-30 degrees C using the giant excised patch technique. The I(NCX) for both isoforms exhibited Na(+)-dependent inactivation and Ca(2+)-dependent positive regulation. The I(NCX) of NCX1.1 exhibited typical mammalian temperature sensitivities with Q(10) values of 2.4 and 2.6 for peak and steady-state currents, respectively. However, the I(NCX) of NCX-TR1.0 was relatively temperature insensitive with Q(10) values of 1.2 and 1.1 for peak and steady-state currents, respectively. I(NCX) current decay was fit with a single exponential, and the resultant rate constant of inactivation (lambda) was determined as a function of temperature. As expected, lambda decreased monotonically with temperature for both isoforms. Although lambda was significantly greater in NCX1.1 compared with NCX-TR1.0 at all temperatures, the effect of temperature on lambda was not different between the two isoforms. These data suggest that the disparities in I(NCX) temperature dependence between these two exchanger isoforms are unlikely due to differences in their inactivation kinetics. In addition, similar differences in temperature dependence were observed in both isoforms after alpha-chymotrypsin treatment that renders the exchanger in a deregulated state. These data suggest that the differences in I(NCX) temperature dependence between the two isoforms are not due to potential disparities in either the I(NCX) regulatory mechanisms or structural differences in the cytoplasmic loop but are likely predicated on differences within the transmembrane segments.