α_1C (Ca_V1.2) L-type calcium channel mediates mechanosensitive calcium regulation

Abstract

Smooth muscle exhibits mechanosensitivity independent of neural input, suggesting that mechanosensitive pathways reside within smooth muscle cells. The native L-type calcium current recorded from human intestinal smooth muscle is modulated by stretch. To define mechanosensitive mechanisms involved in the regulation of smooth muscle calcium entry, we cloned the alpha(1C) L-type calcium channel subunit (Ca(V)1.2) from human intestinal smooth muscle and expressed the channel in a heterologous system. This channel subunit retained mechanosensitivity when expressed alone or coexpressed with a beta(2) calcium channel subunit in HEK-293 or Chinese hamster ovary cells. The heterologously expressed human cardiac alpha(1C) splice form also demonstrated mechanosensitivity. Inhibition of kinase signaling did not affect mechanosensitivity of the native channel. Truncation of the alpha(1C) COOH terminus, which contains an inhibitory domain and a proline-rich domain thought to mediate mechanosensitive signaling from integrins, did not disrupt mechanosensitivity of the expressed channel. These data demonstrate mechanical regulation of calcium entry through molecularly identified L-type calcium channels in mammalian cells and suggest that the mechanosensitivity resides within the pore forming alpha(1C)-subunit.

Characterization of Cav1.4 Complexes (α11.4, β2, and α2δ4) in HEK293T Cells and in the Retina

“…2). A ␤ 2 variant with sequence identical to the retinal ␤ 2X13 was isolated from human jejunum (GenBank TM number AF465485) (41). Although ␤ 2X13 is expressed at higher levels than ␤ 2a in the retina, we were also able to detect the expression of ␤ 2a transcripts that include exon 7A (Fig.…”

“…1 and 55). Of the four classes of Ca v ␤ that have been characterized, ␤ 2 is subject to the most significant alternative splicing, with 13 variants thus far identified (1,41). Despite the evidence in favor of ␤ 2 as a Ca v 1.4 subunit, there has been no characterization of Ca v ␤ 2 variants expressed in human retina.…”

T-type Ca2+channels and autoregulation of local blood flow

“…41,54 However, a direct pressure-dependent activation of Ca 2C influx via VGCCs cannot play a major role in the myogenic response, since myogenic depolarization is dependent on opening of non-selective cation channels, 30,33 and is not affected by application of the L-type antagonist nifedipine. 114,115 The role of Ca V 3.1 channels (and perhaps Cav3.3 channels in human MCAs) in myogenic tone at lower pressures can be explained by a corresponding low myogenic depolarization in the range from approximately ¡65 to ¡50 mV, which activates a non-inactivation window-type Ca 2C current through these channels.…”

“…20,39,40 The Ca V 1.2 a 1 -subunit isoforms cloned from human intestinal smooth muscle and the heart were both shown to be inherently mechano-sensitive when expressed in HEK-293 cells. 41 The Ca V 2 family (HVA) encoding the P/Q-type (Ca V 2.1a/b: splice variants of the Cacna1a gene), Ntype (Ca V 2.2) and R-type (Ca V 2.3) currents are primarily expressed and functional in neurons where they participate in neurotransmitter release, action potentials, and local Ca 2C transients. Several peptide toxins such as v-agatoxins (P/Q-type), some v-conotoxins (N-type) and SNX-482 (R-type) inhibit the Ca V 2 channels specifically and are therefore valuable pharmacological tools (see 34 ).…”

Syntrophin γ2 Regulates SCN5A Gating by a PDZ Domain-mediated Interaction

“…Bath perfusion was used to assess mechanosensitivity as previously described (25,26). The bath was perfused at 10 ml/min for 30 s to create shear stress and activate the mechanosensitive Na ϩ channel according to a previously established protocol (13).…”