Smooth muscle cell proliferation is an important process in arteriogenesis, neointimal hyperplasia and arteriosclerosis (Newby & Zaltsman, 2000). Cellular processes controlling transition from contractile to proliferative phenotype are not well understood, but seem to involve changes in ion channel function or expression (Golovina, 1999). In this study we investigated whether there are differences in Ca²⁺ channels associated with proliferating and growth-arrested phenotypes of smooth muscle cells isolated from human left internal mammary artery.

Discarded arterial samples were obtained anonymously with ethical approval. The medial layer of arteries was removed and digested with collagenase and elastase to release single smooth muscle cells. Cells were maintained in growth culture medium for 6 days and then for a further 2 days with or without growth factors. In growth medium, cells proliferated and adopted a flat kite-like shape. Removal of growth factors arrested proliferation and caused cells to adopt a long, thin spindle-like shape. Both sets of cells labelled with a monoclonal antibody to smooth muscle α-actin. Intracellular Ca²⁺ signals were measured after cells were incubated with 1 µM fura-PE3-AM for 1 h prior to sampling images at 340 and 380 nm excitation wavelengths. In growth-arrested cells, the [Ca²⁺]_i signal on reapplication of 1.5 mM Ca²⁺ to the bath solution was 4.0 times greater (n = 34, P < 0.00001, Student’s t test) if cells were pretreated with 1 µM thapsigargin, suggesting store-operated Ca²⁺ channels were present. Also, an antibody specific for TrpC1 (a store-operated Ca²⁺ channel subunit in vascular myocytes) labelled the plasma membrane. In the presence of 10 µM methoxyverapamil to block L-type voltage-gated Ca²⁺ channels, the [Ca²⁺]_i signal on reapplication of 1.5 mM Ca²⁺ was 2.5 times greater (n = 27, P < 0.00001) in proliferating compared with growth-arrested cells, suggesting enhanced store-operated Ca²⁺ entry. Conversely, there was a significant response to methoxyverapamil in growth-arrested (n = 13) but not proliferating cells (n = 14, P < 0.05). Furthermore, [Ca²⁺]_i was elevated in growth-arrested (n = 4) but not proliferating cells (n = 5) in response to the dihydropyridine agonist (+) 202-791 (0.1-1 µM). Thus functional L-type voltage-gated Ca²⁺ channels appear to be lost from proliferating cells, as suggested by studies on rat aortic myocytes (Quignard et al. 2001). The data support the view that store-operated Ca²⁺ channels are enhanced and L-type voltage-gated Ca²⁺ channels inhibited or lost when human arterial smooth muscle cells are in the proliferative state.

Research funds were from National Heart Research Fund and MRC.

Golovina, V.A. (1999). Am. J. Physiol. 277, C343-349.

Newby, A.C. & Zaltsman, A.B. (2000). J. Pathol. 190, 300-309.

Quignard, J., Harricane, M., Menard, C., Lory, P., Nargeot, J., Capron, L., Mornet, D. & Richard, S. (2001). Cardiovasc. Res. 49, 177-188.