Molecular and Functional Characterization of a Family of Rat Brain T-type Calcium Channels

Abstract

Voltage-gated calcium channels represent a heterogenous family of calcium-selective channels that can be distinguished by their molecular, electrophysiological, and pharmacological characteristics. We report here the molecular cloning and functional expression of three members of the low voltage-activated calcium channel family from rat brain (␣ 1G , ␣ 1H , and ␣ 1I ). Northern blot and reverse transcriptase-polymerase chain reaction analyses show ␣ 1G , ␣ 1H , and ␣ 1I to be expressed throughout the newborn and juvenile rat brain. In contrast, while ␣ 1G and ␣ 1H mRNA are expressed in all regions in adult rat brain, ␣ 1I mRNA expression is restricted to the striatum. Expression of ␣ 1G , ␣ 1H , and ␣ 1I subunits in HEK293 cells resulted in calcium currents with typical T-type channel characteristics: low voltage activation, negative steady-state inactivation, strongly voltage-dependent activation and inactivation, and slow deactivation. In addition, the direct electrophysiological comparison of ␣ 1G , ␣ 1H , and ␣ 1I under identical recording conditions also identified unique characteristics including activation and inactivation kinetics and permeability to divalent cations. Simulation of ␣ 1G , ␣ 1H , and ␣ 1I T-type channels in a thalamic neuron model cell produced unique firing patterns (burst versus tonic) typical of different brain nuclei and suggests that the three channel types make distinct contributions to neuronal physiology.

Induction of T-type Calcium Channel Gene Expression by Chronic Hypoxia

“…As shown in Fig. 1B, a probe specific for the ␣ 1H subunit hybridized to a single ϳ8.5-kb transcript in PC12 cells, which was also detected in rat brain as previously reported (30). A second less abundant band of ϳ10 kb, detected in rat brain, was not present in PC12 cells.…”

“…To our knowledge, T-type Ca 2ϩ channels have not previously been reported in quiescent PC12 cells. We first tested for the presence of mRNAs coding for T-type Ca 2ϩ channel genes by performing reverse transcriptase-PCR experiments using oligonucleotides specific for each of the three previously identified T-type Ca 2ϩ channel isoforms (␣ 1G , ␣ 1H , and ␣ 1I ) (30). Fig.…”

CaV 3.1 and CaV 3.3 account for T-type Ca2+ current in GH3 cells

“…Accordingly, in the present study we measured Ba 2+ currents which were found to be identical to the currents measured when Ca 2+ was used (data not shown). These results allow us to suggest that GH3 cells may not express the α1H isoform, which has been shown to conduct Ba 2+ better than Ca 2+ (10).…”

“…The rate of deactivation was measured as the rate of decay of the tail currents at different repolarizing membrane potentials ( Figure 4 and Table 1). Somewhat surprisingly, the tail currents were best fitted by a double exponential, an unexpected finding since deactivation kinetics described for Ttype Ca 2+ current should follow a monoexponential decay (10). However, there are recent reports suggesting that CaV3.3 (α1I) channels close in a biexponential manner (15)(16)(17).…”

“…Although only 30 ms is shown in Figure 5, the currents were fitted over the entire 100-ms pulse duration. To our surprise, the fast and slow time constants measured in fluoride-rich internal solution showed previous sequencing data for the α1G and α1I subunits (10). We also used primers specific for ß-actin as a positive control (Figure 6, lanes 2 to 4).…”