Electrical Membrane Activity and Intracellular Calcium Buffering Control Exocytosis Efficiency in <i>Xenopus</i> Melanotrope Cells

Abstract

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.

Neuronal, Neurohormonal, and Autocrine Control of Xenopus Melanotrope Cell Activity

“…These steps are generated by electrical plasma membrane bursting activity (action currents) that induce Ca 2+ influx through voltage-operated plasma membrane Ca 2+ channels (VOCCs). [60][61][62][63][64][65][66] We showed in Xenopus melanotropes that activation of the CaR by the selective CaR agonist, L-phenylalanine, stimulates the frequency of the Ca 2+ oscillations, which may underlie the autoexcitatory stimulation by the CaR of αMSH release. 59 Whether the CaR acts by controlling action currents is currently under investigation.…”

“…[67][68][69] The Ca 2+ oscillations are generated by Ca 2+ influx through VOCCs during bursts of action potential firing, followed by wave propagation through the cell via Ca 2+ -induced Ca 2+ release. 70 Ca 2+ influx during the bursts of action potentials sets the level of release activity 63 in a way that may be dynamically controlled by the bursting frequency. 71 Some aspects of this pattern with slow kinetics may be important for the regulation of POMC precursor gene expression.…”

Dopamine D₂-Receptor Activation Differentially Inhibits N- and R-Type Ca²⁺ Channels in <i>Xenopus</i> Melanotrope Cells

“…For N-type channels the reversal of the β/γ-induced inhibition requires much shorter pre-pulse durations [18]. Therefore, apomorphin-induced inhibition of the I Ca was also checked when the duration of pre-pulse was reduced to 10 ms and the interval time between the second pulse and pre-pulse was 1 ms (fig.…”

“…Cells were prepared as described previously [18]. In short, after anesthetization in tap water containing 0.1% (w/v) MS222 and 20 m M NaHCO 3 , animals were perfused with Xenopus Ringer’s solution (112 m M NaCl, 2 m M KCl, 2 m M CaCl 2 , 15 m M Hepes, 10 m M glucose, 0.025% MS222, pH 7.4) containing 0.025% (w/v) MS222, to remove blood cells.…”

Melanotrope Cells of Xenopus laevis Express Multiple Types of High‐Voltage‐Activated Ca2+ Channels

“…Cell preparations were made as described previously (19). In brief, after anaesthetisation, animals were perfused with Xenopus Ringer's solution (112 m M NaCl, 2 m M KCl, 2 m M CaCl 2 , 15 m M HEPES, 10 m M glucose; pH 7.4), containing 0.025% (w/v) MS222 (Sigma, St Louis, MO, USA), to remove blood cells.…”