Ion currents regulated by acute and chronic osmotic stimuli in rat supraoptic nucleus neurons
The magnocellular neurosecretory cells (MNCs) of the hypothalamus are able to change their firing rate and pattern in response to small changes in external osmolality due to the involvement of osmosensitive ion channels. The firing rate and pattern determine the release of vasopressin (VP), a primary hormone regulating osmolality by controlling water excretion from the kidney. Both VP- and oxytocin (OT)-MNCs display irregular and infrequent fire when plasma osmolality is near normal, and they progressively increase the frequency of firing to fast continuous firing with increases in osmolality. VP-MNCs also respond to osmotic stimulation by adopting a phasic pattern of firing, which maximizes neuropeptide secretion. Sustained dehydration also causes structural and functional adaptations in MNCs. Voltage-dependent Ca2+ channels play many important roles not only in the regulation of cell excitability but also in intracellular signal transduction, and L-type Ca2+ channel-mediated Ca2+ signals initiate intracellular signal transduction events that activate long-lasting changes in brain function and behavior. Our electrophysiological and immunocytochemical studies demonstrate that 16-24 h of water deprivation causes a significant increase in the amplitude of L-type Ca2+ current (from –55.5 ± 6.2 to –99.1 ± 10.0 pA) but not in other types of Ca2+ current. This increase occurred in both VP- and OT-MNCs. Such an increase in L-type Ca2+ current may contribute to modulation of firing rate and pattern, regulation of vasopressin release, structural adaptation in MNCs during sustained dehydration. The mechanisms underlying the transition of the electrical behaviour are not completely understood. Ion channels, especially osmosensitive ion channels, play key roles in the modulation of MNC firing. A voltage-gated, 4-AP- and TEA-insensitive slowly activating outward current displayed a significant increase in about 66% of MNCs when the osmolality of the external solution was acutely increased from 295 to 325 mosmol kg-1. The responding cells showed an increase in net outward current from 12.3 ± 1.3 pA/pF to 21.4 ± 1.8 pA/pF. The reversal potential of this current was near the equilibrium for K+ and shifted with changes of K+ concentrations in external solution, suggesting that this current is a K+-selective current. The KCNQ/M current selective blockers linopirdine (150 µM) and XE991 (5 µM) suppressed this current. The IC50 of XE991 blockade was 3.9 ìM. The KCNQ/M channel openers retigabine (10 µM) and flupirtine (10 µM) significantly increased the current and shifted its activation curve toward more negative potentials. E4031, a specific blocker of ERG K+ channels, did not significantly block this current. The results from immunocytochemistry suggest that MNCs express KCNQ2, KCNQ3, KCNQ4, and KCNQ5, but not KCNQ1. These data suggest that this osmosensitive current could be a KCNQ/M current. Studies using single unit extracellular recording in hypothalamic explants showed that 10 µM XE991 increased MNC firing rate and that 20 µM retigabine decreased firing rate or caused a cessation of firing. These data suggest that a KCNQ/M current contributes to the regulation of MNC firing. KCNQ/M channels play key roles in regulating neuronal excitability in many types of central neurons. Slow activation of this current during firing might suppress activity by hyperpolarizing the cells and thus contribute to a transition between fast continuous and burst firing. Our studies will be beneficial to understand the mechanisms that control VP and OT in response to acute changes in osmolality and also the mechanisms underlying MNC adaptation during sustained dehydration.
DegreeDoctor of Philosophy (Ph.D.)
SupervisorFisher, Thomas E.
CommitteeWalz, Wolfgang; Howland, John; Ferguson, Alastair; Desautels, Michel; West, Nigel
L-type Ca2+ channel