Abstract
Transmembrane Ca2+ influx is essential to the proper functioning of the central clock in the suprachiasmatic nucleus (SCN). In the rat SCN neurons, the clearance of somatic Ca2+ following depolarization-induced Ca2+ transients involves Ca2+ extrusion via Na+/Ca2+ exchanger (NCX) and mitochondrial Ca2+ buffering. Here we show an important role of intracellular Na+ in the regulation of [Ca2+]i in these neurons. The effect of Na+ loading on [Ca2+]i was determined with the Na+ ionophore monensin and the cardiac glycoside ouabain to block Na+/K+-ATPase (NKA). Ratiometric Na+ and Ca2+ imaging was used to measure the change in [Na+]i and [Ca2+]i, and cell-attached recordings to investigate the effects of monensin and ouabain on spontaneous firing. Our results show that in spite of opposite effects on spontaneous firing and basal [Ca2+], both monensin and ouabain induced Na+ loading, and increased the peak amplitude, slowed the fast decay rate, and enhanced the slow decay phase of 20 mM K+-evoked Ca2+ transients. Furthermore, both ouabain and monensin preferentially enhanced nimodipine-insensitive Ca2+ transients. Together, our results indicate that in the SCN neurons the NKA plays an important role in regulating [Ca2+]i, in particular, associated with nimodipine-insensitive Ca2+ channels.
Highlights
The central clock in the hypothalamic suprachiasmatic nucleus (SCN) controls circadian rhythms in mammals [1]
Intracellular Ca2+ is essential to the proper functioning of the SCN, and its homeostasis depends on various Ca2+ handling systems, including those involved in mediating Ca2+ entry, extrusion (plasmalemmal Na+/Ca2+ exchanger (NCX) and Ca2+-ATPase), and buffering (Ca2+ binding proteins, endoplasmic reticulum, and mitochondria)
To investigate the regulation of Ca2+ homeostasis in the SCN neurons, we recently reported that the SCN expresses NCX1 and NCX2, with NCX1 distributed in the whole SCN, and NCX2 restricted to the retinorecipient ventral SCN [10]
Summary
The central clock in the hypothalamic suprachiasmatic nucleus (SCN) controls circadian rhythms in mammals [1]. Superimposition of the Ca2+ transients in ouabain indicates an increase in the amplitude and a slowing in the fast decay rate from 5 (trace b), 10 (trace c), to 15 (trace d) min and leveling off thereafter to 35 (trace h) min into the application of ouabain (Figure 5C) This is best seen by superimposing the normalized Ca2+. For the first cell (top), after an initial increase in ouabain the basal level of [Ca2+]i returned to near its resting level for as long as 30 min along with the appearance of prominent slow decay phase (marked by arrowheads) for the 20 K+-evoked Ca2+ transients.
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