Entry In Smooth Muscle Research Articles

Thapsigargin decreases the Na+- Ca2 + exchanger mediated Ca2 + entry in pig coronary artery smooth muscle

Na+- Ca2+ exchanger (NCX) has been proposed to play a role in refilling the sarco/endoplasmic reticulum (SER) Ca2+ pool along with the SER Ca2+ pump (SERCA). Here, SERCA inhibitor thapsigargin was used to determine the effects of SER Ca2+ depletion on NCX–SERCA interactions in smooth muscle cells cultured from pig coronary artery. The cells were Na+-loaded and then placed in either a Na+-containing or in a Na+-substituted solution. Subsequently, the difference in Ca2+ entry between the two groups was examined and defined as the NCX mediated Ca2+ entry. The NCX mediated Ca2+ entry in the smooth muscle cells was monitored using two methods: Ca2+sensitive fluorescence dye Fluo-4 and radioactive Ca2+. Ca2+-entry was greater in the Na+-substituted cells than in the Na+-containing cells when measured by either method. This difference was established to be NCX-mediated as it was sensitive to the NCX inhibitors. Thapsigargin diminished the NCX mediated Ca2+ entry as determined by either method. Immunofluorescence confocal microscopy was used to determine the co-localization of NCX1 and subsarcolemmal SERCA2 in the cells incubated in the Na+-substituted solution with or without thapsigargin. SER Ca2+ depletion with thapsigargin increased the co-localization between NCX1 and the subsarcolemmal SERCA2. Thus, inhibition of SERCA2 leads to blockade of constant Ca2+ entry through NCX1 and also increases proximity between NCX1 and SERCA2. This blockade of Ca2+ entry may protect the cells against Ca2+-overload during ischemia–reperfusion when SERCA2 is known to be damaged.

A novel receptor-operated Ca2+-permeable channel activated by ATP in smooth muscle

Receptor-operated Ca2+ entry has been proposed as a signalling mechanism in many cells. Receptor-operated Ca2+ channels (ROCs) were first postulated in smooth muscle by Bolton, van Breemen and Somlyo and Somlyo, but recordings of directly ligand-gated Ca2+ current are lacking. Here we describe receptor-operated Ca2+ current evoked in arterial smooth muscle cells by ATP, a sympathetic neurotransmitter. ATP activates channels with approximately 3:1 selectivity for Ca2+ over Na+ at near-physiological concentrations and with a unitary conductance of approximately 5 pS in 110 mM Ca2+ or Ba2+. The channels can be opened even at very negative potentials and resist inhibition by cadmium or nifedipine, unlike voltage-gated Ca2+ channels; they are not blocked by Mg2+, unlike NMDA (N-methyl-D-aspartate)-activated channels; they are directly activated by ligand, without involvement of readily diffusible second messengers, unlike cation channels in neutrophils and T lymphocytes. Thus, the ATP-activated channels provide a distinct mechanism for excitatory synaptic current and Ca2+ entry in smooth muscle.