Effects of temperature on human L-type cardiac Ca 2+ channels expressed in Xenopus oocytes

Abstract

Temperature normally affects peak L-type Ca2+ channel (CaCh) current with a temperature coefficient (Q10) of between 1.8 and 3.5; in cardiomyocytes attenuating protein kinase A activity increases Q10 whilst activating it lowers Q10. We examine temperature effects using cloned human cardiac CaChs expressed in Xenopus oocytes. Peak inward currents (IBa) through expressed CaChs (i.e. alpha1C alpha2/deltaa beta1b) exhibited a Q10 of 5.8+/-0.4 when examined between 15 and 25 degreesC. The nifedipine-sensitive IBa exhibited a higher Q10 of 8.7+/-0.5, whilst the nifedipine-insensitive IBa exhibited Q10 of 3.7+/-0.3. Current/voltage (I/V) relationships shifted to negative potentials on warming. Using instead a different CaCh beta subunit isoform, beta2c, gave rise to an IBa similar to those expressed using beta1b. We utilized a carboxyl deletion mutant, alpha1C-Delta1633, to determine the temperature sensitivity of the pore moiety in the absence of auxiliary subunits; IBa through this channel exhibited a Q10 of 9.3+/-0.3. However, the Q10 for macroscopic conductance was reduced compared to that of heteromeric channels; decreasing from 5.0 (i.e. alpha1C alpha2/deltaa beta1b) and 3.9 (i.e. alpha1C alpha2/deltaa beta2c) to 2.4 (alpha1C-Delta1633). These observations differ markedly from those made in studies of cardiomyocytes, and suggest that enhanced sensitivity may depend on the membrane environment, channel assembly or other regulatory factors.

Cloning of a Functional Splice Variant of L-type Calcium Channel β2 Subunit from Rat Heart

“…Our findings also confirmed that the short N-terminal region was important for modification of inactivation kinetics, because ␤ 2a and ␤ 2c subunits were only different in this region. The N-terminal region of the rat ␤ 2c subunit was identical to those of a Lambert-Eaton myasthenic syndrome antigen and the human ␤ 2c subunit, except for some amino acids considered to be species differences (23,30). The similarity to the former protein raises the possibility that the rat ␤ 2c subunit was cloned from neuron, which innervated into heart.…”

“…No consensus site for cAMP-dependent protein kinase was found in this region. In contrast to other ␤ 2 subunits, the sequence for the N-terminal region of the human ␤ 2c subunit, which was also cloned from human ventricular septum (23), is almost identical to that for the rat ␤ 2c subunit.…”

“…2B, along with that for the human ␤ 2c , the rabbit ␤ 2a , and the rabbit ␤ 2b subunits (7,8,23). The rat ␤ 2c subunit differs only in the N-terminal region from the rat ␤ 2a , the rabbit ␤ 2a , and ␤ 2b subunits except for some amino acids sequence, considered to be species differences.…”

“…Among them, the N-terminal region of the rat ␤ 2c subunit was identical to the human ␤ 2c subunit except for some differences considered to be species-specific. However, the I-V relationship and the inactivation kinetics of the human ␤ 2c subunit reported were quite different from those of the rat ␤ 2c subunit (23). These differences may be caused by variations in the experimental conditions because Allen et al (23) used Xenopus oocyte as the heterologous expression system and recorded I Ba at 15, 20, and 25°C, using 40 mM Ba 2ϩ as charge carrier.…”

The genetic basis of Brugada syndrome: A mutation update

“…CACNB2 consists of 14 exons covering 421 kb at chromosome 10p12 [Allen and Mikala, 1998;Taviaux et al, 1997]. Ca v b2 functions as a chaperone for the a-subunit of Ca v 1.2, ensuring its transport to the plasma membrane [Cornet et al, 2002].…”