Regulation and function of Cav3.1 T-type calcium channels in IGF-I-stimulated pulmonary artery smooth muscle cells

Arterial smooth muscle cells enter the cell cycle and proliferate in conditions of disease and injury, leading to adverse vessel remodeling. In the pulmonary vasculature, diverse stimuli cause proliferation of pulmonary artery smooth muscle cells (PASMCs), pulmonary artery remodeling, and the clinical condition of pulmonary hypertension associated with significant health consequences. PASMC proliferation requires extracellular Ca2+ influx that is intimately linked with intracellular Ca2+ homeostasis. Among the primary sources of Ca2+ influx in PASMCs is the low-voltage-activated family of T-type Ca2+ channels; however, up to now, mechanisms for the action of T-type channels in vascular smooth muscle cell proliferation have not been addressed. The Cav3.1 T-type Ca2+ channel mRNA is upregulated in cultured PASMCs stimulated to proliferate with insulin-like growth factor-I (IGF-I), and this upregulation depends on phosphatidylinositol 3-kinase/Akt signaling. Multiple stimuli that trigger an acute rise in intracellular Ca2+ in PASMCs, including IGF-I, also require the expression of Cav3.1 Ca2+ channels for their action. IGF-I also led to cell cycle initiation and proliferation of PASMCs, and, when expression of the Cav3.1 Ca2+ channel was knocked down by RNA interference, so were the expression and activation of cyclin D, which are necessary steps for cell cycle progression. These results confirm the importance of T-type Ca2+ channels in proper progression of the cell cycle in PASMCs stimulated to proliferate by IGF-I and suggest that Ca2+ entry through Cav3.1 T-type channels in particular interacts with Ca2+-dependent steps of the mitogenic signaling cascade as a central component of vascular remodeling in disease.

Download Full-text

Diversity of voltage-dependent K+ channels in human pulmonary artery smooth muscle cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00438.2003 ◽

2004 ◽

Vol 287 (1) ◽

pp. L226-L238 ◽

Cited By ~ 41

Author(s):

Oleksandr Platoshyn ◽

Carmelle V. Remillard ◽

Ivana Fantozzi ◽

Mehran Mandegar ◽

Tiffany T. Sison ◽

...

Keyword(s):

Smooth Muscle ◽

Pulmonary Artery ◽

Smooth Muscle Cells ◽

Critical Role ◽

Muscle Cells ◽

Pulmonary Vasculature ◽

Wide Range ◽

Voltage Dependent ◽

Pulmonary Artery Smooth Muscle ◽

Α Subunits

Electrical excitability, which plays an important role in excitation-contraction coupling in the pulmonary vasculature, is regulated by transmembrane ion flux in pulmonary artery smooth muscle cells (PASMC). This study examined the heterogeneous nature of native voltage-dependent K+ channels in human PASMC. Both voltage-gated K+ (KV) currents and Ca2+-activated K+ (KCa) currents were observed and characterized. In cell-attached patches of PASMC bathed in Ca2+-containing solutions, depolarization elicited a wide range of K+ unitary conductances (6–290 pS). When cells were dialyzed with Ca2+-free and K+-containing solutions, depolarization elicited four components of KV currents in PASMC based on the kinetics of current activation and inactivation. Using RT-PCR, we detected transcripts of 1) 22 KV channel α-subunits (KV1.1–1.7, KV1.10, KV2.1, KV3.1, KV3.3–3.4, KV4.1–4.2, KV5.1, KV 6.1–6.3, KV9.1, KV9.3, KV10.1, and KV11.1), 2) three KV channel β-subunits (KVβ1–3), 3) four KCa channel α-subunits ( Slo-α1 and SK2–SK4), and 4) four KCa channel β-subunits (KCaβ1–4). Our results show that human PASMC exhibit a variety of voltage-dependent K+ currents with variable kinetics and conductances, which may result from various unique combinations of α- and β-subunits forming the native channels. Functional expression of these channels plays a critical role in the regulation of membrane potential, cytoplasmic Ca2+, and pulmonary vasomotor tone.

Download Full-text

Endothelin-1 increases intracellular Ca2+ in rabbit pulmonary artery smooth muscle cells through phospholipase C

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00131.2005 ◽

2005 ◽

Vol 289 (4) ◽

pp. H1551-H1559 ◽

Cited By ~ 15

Author(s):

Eun A. Ko ◽

Won Sun Park ◽

Jae-Hong Ko ◽

Jin Han ◽

Nari Kim ◽

...

Keyword(s):

Smooth Muscle ◽

Pulmonary Artery ◽

Smooth Muscle Cells ◽

Phospholipase C ◽

Muscle Cells ◽

Transient Increase ◽

Rabbit Pulmonary Artery ◽

Intracellular Ca ◽

Pulmonary Artery Smooth Muscle ◽

Sustained Increase

In freshly isolated rabbit pulmonary artery smooth muscle cells, endothelin (ET)-1 induced a transient increase in intracellular Ca2+ concentration ([Ca2+]i) followed by a return to the initial [Ca2+]i. This response was not abolished by the voltage-dependent Ca2+ channel blocker nicardipine or removal of Ca2+ from the bath solution but was inhibited by ryanodine and thapsigargin. This finding suggested that the increase in [Ca2+]i induced by ET-1 was attributable to release of Ca2+ from ryanodine- and inositol 1,4,5-trisphosphate-sensitive intracellular Ca2+ stores. The transient increase in [Ca2+]i induced by ET-1 was also inhibited by pretreatment with antagonists of ET type A and B (ETA and ETB) receptors (BQ-123 and BQ-788, respectively). Furthermore, the ETB receptor agonist IRL-1620 induced an increase in [Ca2+]i that was followed by a sustained increase in [Ca2+]i; the sustained increase in [Ca2+]i was blocked by nicardipine. Using the nystatin-perforated patch-clamp technique, we found that IRL-1620 caused an increase in Ca2+ current that was inhibited by addition of ET-1. ET-1 did not inhibit Ca2+ current when cells were pretreated with BQ-123. These results suggested that when both receptor types are activated, the opposing responses lead to abolition of the sustained [Ca2+]i increases induced by ETB receptor activation. Western blot analysis confirmed expression of ETA and ETB receptors. Finally, U-73122 inhibited the ET-1-induced [Ca2+]i increase, indicating that phospholipase C was involved in modulation of the ET-1-induced [Ca2+]i increase in rabbit pulmonary artery smooth muscle cells.

Download Full-text

Heterogeneous activation of p19Arf in pulmonary artery smooth muscle cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00117.2010 ◽

2011 ◽

Vol 300 (4) ◽

pp. L642-L647

Author(s):

Victor Solodushko ◽

Diego F. Alvarez ◽

Ryan Viator ◽

Tiffany Messerall ◽

Brian Fouty

Keyword(s):

Cell Cycle ◽

Smooth Muscle ◽

Pulmonary Artery ◽

Smooth Muscle Cells ◽

Time Course ◽

Muscle Cells ◽

Tumor Formation ◽

Wild Type ◽

Daughter Cells ◽

Pulmonary Artery Smooth Muscle

p19ARF is a tumor suppressor that leads to cell cycle arrest or apoptosis by stabilizing p53. p19ARF is not critical for cell cycle regulation under normal conditions, but loss of p19ARF is seen in many human cancers, and a murine p19Arf knockout model leads to malignant proliferation and tumor formation; its role in controlling nonmalignant proliferation is less defined. To examine this question, pulmonary artery smooth muscle cells (PASMC) were expanded in culture from a transgenic mouse in which the coding sequence of the p19Arf gene was replaced with a cDNA encoding green fluorescent protein (GFP), leaving the promoter intact. During the first 10 days in culture, wild-type, heterozygous, and knockout PASMC grew similarly, but, by day 14, p19Arf-deficient PASMC proliferated faster than p19Arf heterozygous or wild-type cells; reexpression of p19Arf prevented the increased proliferation. This time course correlated with activation of the p19Arf promoter, as indicated by the appearance of GFP positivity in p19Arf-deficient PASMC. By day 42, ∼80% of p19Arf-deficient cells were GFP-positive. When GFP-positive, p19Arf-deficient cells were sorted and subcultured separately, they remained GFP-positive, indicating that once cells had activated the p19Arf promoter, the promoter remained active in those and all subsequent daughter cells. In contrast, GFP-negative p19Arf-deficient cells gave rise to a combination of GFP-positive and -negative daughter cells over time. These results suggest that a subpopulation of PASMC are resistant to the signals that activate the p19Arf promoter, an event that would normally target these cells for arrest or cell death.

Download Full-text

Silencing Of NHE1 Gene Decreases Hypoxia-induced Proliferation And Cell Cycle Progression Of Human Pulmonary Artery Smooth Muscle Cells

10.1164/ajrccm-conference.2010.181.1_meetingabstracts.a3950 ◽

2010 ◽

Author(s):

Lunyin Yu ◽

Charles A. Hales

Keyword(s):

Cell Cycle ◽

Smooth Muscle ◽

Pulmonary Artery ◽

Smooth Muscle Cells ◽

Cell Cycle Progression ◽

Muscle Cells ◽

Cycle Progression ◽

Pulmonary Artery Smooth Muscle ◽

Human Pulmonary Artery

Download Full-text

Dexfenfluramine increases pulmonary artery smooth muscle intracellular Ca2+, independent of membrane potential

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.1999.277.3.l662 ◽

1999 ◽

Vol 277 (3) ◽

pp. L662-L666 ◽

Cited By ~ 15

Author(s):

Helen L. Reeve ◽

Stephen L. Archer ◽

Marjorie Soper ◽

E. Kenneth Weir

Keyword(s):

Smooth Muscle ◽

Membrane Potential ◽

Pulmonary Artery ◽

Smooth Muscle Cells ◽

Primary Pulmonary Hypertension ◽

Muscle Cells ◽

Channel Activity ◽

Intracellular Ca ◽

Pulmonary Artery Smooth Muscle ◽

Dose Dependent

The anorexic agent dexfenfluramine causes the development of primary pulmonary hypertension in susceptible patients by an unknown mechanism that may include changes in K+-channel activity and intracellular Ca2+ concentration ([Ca2+]i). We investigated the dose-dependent effects of dexfenfluramine on [Ca2+]i, K+ current, and membrane potential in freshly dispersed rat pulmonary artery smooth muscle cells. Dexfenfluramine caused a dose-dependent (1–1,000 μM) increase in [Ca2+]i, even at concentrations lower than those necessary to inhibit K+ currents (10 μM) and cause membrane depolarization (100 μM). The [Ca2+]iresponse to 1 and 10 μM dexfenfluramine was completely abolished by pretreatment of the cells with 0.1 μM thapsigargin, whereas the response to 100 μM dexfenfluramine was reduced. CoCl2 (1 mM), removal of extracellular Ca2+, and pretreatment with caffeine (1 mM) reduced but did not abolish the response to 100 μM dexfenfluramine. We conclude that dexfenfluramine increases [Ca2+]iin rat pulmonary artery smooth muscle cells by both release of Ca2+ from the sarcoplasmic reticulum and influx of extracellular Ca2+.

Download Full-text