Cholecystokinin Activates Orexin/Hypocretin Neurons through the Cholecystokinin A Receptor

Abstract

Orexin A and B are neuropeptides implicated in the regulation of sleep/wakefulness and energy homeostasis. The regulatory mechanism of the activity of orexin neurons is not precisely understood. Using transgenic mice in which orexin neurons specifically express yellow cameleon 2.1, we screened for factors that affect the activity of orexin neurons (a total of 21 peptides and six other factors were examined) and found that a sulfated octapeptide form of cholecystokinin (CCK-8S), neurotensin, oxytocin, and vasopressin activate orexin neurons. The mechanisms that underlie CCK-8S-induced activation of orexin neurons were studied by both calcium imaging and slice patch-clamp recording. CCK-8S induced inward current in the orexin neurons. The CCK A receptor antagonist lorglumide inhibited CCK-8S-induced activation of orexin neurons, whereas the CCK B receptor agonists CCK-4 (a tetrapeptide form of cholecystokinin) and nonsulfated CCK-8 had little effect. The CCK-8S-induced increase in intracellular calcium concentration was eliminated by removing extracellular calcium but not by an addition of thapsigargin. Nifedipine, -conotoxin, -agatoxin, 4-ethylphenylamino-1,2-dimethyl-6-methylaminopyrimidinium chloride, and SNX-482 had little effect, but La 3ϩ , Gd 3ϩ , and 2-aminoethoxydiphenylborate inhibited CCK-8S-induced calcium influx. Additionally, the CCK-8S-induced inward current was dramatically enhanced in the calcium-free solution and was inhibited by the cation channel blocker SKF96365, suggesting an involvement of extracellular calcium-sensitive cation channels. CCK-8S did not induce an increase in intracellular calcium concentration when membrane potential was clamped at Ϫ60 mV, suggesting that the calcium increase is induced by depolarization. The evidence presented here expands our understanding of the regulation of orexin neurons and the physiological role of CCK in the CNS.

Selective loss of GABA B receptors in orexin-producing neurons results in disrupted sleep/wakefulness architecture

“…4 C and D). Moreover, we found that effects of other inhibitory or excitatory neurotransmitters, 5-hydroxytryptamine (5-HT) (19) or cholecystokinin (CCK) (20) were also decreased (Fig. 4 E and F).…”

Vasopressin Increases Locomotion through a V1a Receptor in Orexin/Hypocretin Neurons: Implications for Water Homeostasis

“…However, there is a difference in the source of calcium mobilization between the AVPinduced and the CCK-8S-induced responses. The CCK-8S-induced increase in [Ca 2ϩ ] i was not inhibited by thapsigargin ( Tsujino et al, 2005). In contrast, the AVP-induced response was sensitive to both thapsigargin and xestospongin C. The AVPinduced increase in [Ca 2ϩ ] i was inhibited ϳ50% of control value by these compounds.…”

“…Orexin neurons receive abundant inputs from the limbic system, the raphe nucleus, and the basal forebrain Yoshida et al, 2006). Intracellular calcium in orexin neu-rons is altered by a number of neuroactive substances, including cholecystokinin, neurotensin, AVP, oxytocin, neuropeptide Y (NPY), adenosine, serotonin, and noradrenaline (Fu et al, 2004;Tsujino et al, 2005;Yamanaka et al, 2006;Liu and Gao, 2007).…”

Orexin Directly Excites Orexin Neurons through Orexin 2 Receptor

“…The activation of phospholipase C by the ␤␥ subunit of the G-protein might be involved because OX2R is believed to couple with the Gq and Gi types of G-proteins (Zhu et al, 2003). We previously showed that orexin neurons are activated by CCK and AVP via the G-protein-coupled receptors (Gq), CCK A receptor, and the V1a receptor (Tsujino et al, 2005;Tsunematsu et al, 2008). Both CCK-and AVP-induced inward currents were robustly enhanced by removing the calcium ions from the extracellular solution.…”