In many biological systems, cells display spontaneous calcium oscillations (CaOs) and repetitive action-potential firing. These phenomena have been described separately by models for intracellular inositol trisphosphate (IP3)-mediated CaOs and for plasma membrane excitability. In this study, we present an integrated model that combines an excitable membrane with an IP3-mediated intracellular calcium oscillator. The IP3 receptor is described as an endoplasmic reticulum (ER) calcium channel with open and close probabilities that depend on the cytoplasmic concentration of IP3 and Ca2+. We show that simply combining this ER model for intracellular CaOs with a model for membrane excitability of normal rat kidney (NRK) fibroblasts leads to instability of intracellular calcium dynamics. To ensure stable long-term periodic firing of action potentials and CaOs, it is essential to incorporate calcium transporters controlled by feedback of the ER store filling, for example, store-operated calcium channels in the plasma membrane. For low IP3 concentrations, our integrated NRK cell model is at rest at -70 mV. For higher IP3 concentrations, the CaOs become activated and trigger repetitive firing of action potentials. At high IP3 concentrations, the basal intracellular calcium concentration becomes elevated and the cell is depolarized near -20 mV. These predictions are in agreement with the different proliferative states of cultures of NRK fibroblasts. We postulate that the stabilizing role of calcium channels and/or other calcium transporters controlled by feedback from the ER store is essential for any cell in which calcium signaling by intracellular CaOs involves both ER and plasma membrane calcium fluxes.
In neural and neuroendocrine cells, Ca2+ influx is essential for exocytosis. Ca2+ influx takes place through electrical membrane activity, which often occurs in bursts of action potentials that lead to intracellular Ca2+ oscillations. Cytoplasmic Ca2+ buffers and intracellular Ca2+ stores are involved in the propagation of the oscillations through the cell. Studies focused on action potential bursts with a high frequency up to 20 Hz indicate that, depending on the cell type under investigation, bursts either enhance or reduce exocytosis efficiency. In many cell types, the bursting frequency can be as low as 1 Hz, although no information is present on whether this influences exocytosis efficiency. The present study addresses the role of low-frequency bursts around 1 Hz and cytoplasmic Ca2+ buffering in the regulation of exocytosis efficiency, using neuroendocrine melanotrope cells of the amphibian Xenopus laevis. Exocytosis efficiency was determined by membrane capacitance measurements. Mimicking the bursting activity of 1 Hz (typical for this cell type) by repetitive depolarizing pulses enhanced exocytosis efficiency by 58% compared to application of only one single depolarizing pulse. This increase appears to be particularly due to a small number of distinct depolarizing pulses within a burst. Including the fast Ca2+ buffer BAPTA in the intracellular solution reduced exocytosis efficiency by 60% in the first part of a burst, whereas during the later part of the burst, stimulation (+50%) took place. We conclude that low-frequency bursting in the Xenopus melanotrope cell strongly promotes exocytosis efficiency and that this efficiency also depends on the capacity of the cytoplasm to buffer the intracellular Ca2+ signal; strong Ca2+ buffering during a short burst will decrease exocytosis efficiency, whereas with prolonged bursts, buffering capacity will be overcome, leading to Ca2+ accumulation and thus enhanced exocytosis efficiency.
Cultures of normal rat kidney (NRK) fibroblasts may display spontaneous calcium action potentials which propagate throughout the cellular monolayer. Pacemaking activity of NRK cells was studied by patch clamp electrophysiology and vital calcium imaging, using a new experimental approach in which a ring was placed on the monolayer in order to physically separate pacemakers within or under the ring and follower cells outside the ring. Stimulation of cells inside the ring with IP(3)-generating hormones such as prostaglandin F(2alpha) (PGF(2alpha)) resulted in the induction of periodic action potentials outside the ring, which were abolished when the L-type calcium channel blocker nifedipine was added outside the ring, but not inside the ring. PGF(2alpha)-treated cells displayed asynchronous IP(3)-mediated calcium oscillations of variable frequency, while follower cells outside the ring showed synchronous calcium transients which coincided with the propagating action potential. Mathematical modelling indicated that addition of PGF(2alpha) inside the ring induced both a membrane potential gradient and an intracellular IP(3) gradient, both of which are essential for the induction of pacemaking activity under the ring. These data show that intercellular coupling between PGF(2alpha)-treated and non-treated cells is essential for the generation of a functional pacemaker area whereby synchronization of calcium oscillations occurs by activation of L-type calcium channels.
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