Concentration-dependent activation of dopamine receptors differentially modulates GABA release onto orexin neurons.

Dopamine (DA) and orexin neurons play important roles in reward and food intake. There are anatomical and functional connections between these two cell groups: orexin peptides stimulate DA neurons in the ventral tegmental area and DA inhibits orexin neurons in the hypothalamus. However, the cellular mechanisms underlying the action of DA on orexin neurons remain incompletely understood. Therefore, the effect of DA on inhibitory transmission to orexin neurons was investigated in rat brain slices using the whole-cell patch-clamp technique. We found that DA modulated the frequency of spontaneous and miniature IPSCs (mIPSCs) in a concentration-dependent bidirectional manner. Low (1 µM) and high (100 µM) concentrations of DA decreased and increased IPSC frequency, respectively. These effects did not accompany a change in mIPSC amplitude and persisted in the presence of G-protein signaling inhibitor GDPßS in the pipette, suggesting that DA acts presynaptically. The decrease in mIPSC frequency was mediated by D2 receptors whereas the increase required co-activation of D1 and D2 receptors and subsequent activation of phospholipase C. In summary, our results suggest that DA has complex effects on GABAergic transmission to orexin neurons, involving cooperation of multiple receptor subtypes. The direction of dopaminergic influence on orexin neurons is dependent on the level of DA in the hypothalamus. At low levels DA disinhibits orexin neurons whereas at high levels it facilitates GABA release, which may act as negative feedback to curb the excitatory orexinergic output to DA neurons. These mechanisms may have implications for consummatory and motivated behaviours.

Dopamine acts as a partial agonist for α2A adrenoceptor in melanin-concentrating hormone neurons.

Melanin-concentrating hormone (MCH) is a hypothalamic neuropeptide that promotes positive energy balance and anxiety. Since dopamine (DA) is also closely implicated in these functions, the present study investigated the effect of DA on MCH neurons. Using whole-cell patch-clamp recordings in rat brain slices, we found that DA hyperpolarizes MCH neurons by activating G-protein-activated inwardly rectifying K(+) (GIRK) channels. Pharmacological study indicated that the effect was mediated by α2A adrenoceptors, not DA receptors. DA-induced outward current was also observed in the presence of tetrodotoxin or the dopamine ß-hydroxylase inhibitor fusaric acid, suggesting that DA directly binds to α2A receptors on MCH neurons, rather than acting presynaptically or being transformed into norepinephrine (NE) in the slice preparation. The effects of NE and DA were concentration-dependent with EC(50) of 5.9 and 23.7 µm, respectively, and a maximal effect of 106.6 and 57.2 pA, respectively, suggesting that DA functions as a partial agonist. Prolonged (5 min) activation of α2A receptors by either DA or NE attenuated the subsequent response to DA or NE, while 5 s applications were not sufficient to induce desensitization. Therefore, a history of α2A receptor activation by DA or NE can have a lasting inhibitory effect on the catecholaminergic transmission to MCH neurons. Our study suggests that α2A receptors expressed by MCH neurons may be one of the pathways by which DA and NE can interact and modulate mood and energy homeostasis, and this cross talk may have functional implications in mood disorders and obesity.

Carbonic anhydrase related protein 8 mutation results in aberrant synaptic morphology and excitatory synaptic function in the cerebellum.

Carbonic anhydrase related protein 8 (Car8) is known to be abundantly expressed in Purkinje cells (PCs), and its genetic mutation causes a motor coordination defect. To determine the underlying mechanism, we analyzed the mouse cerebellum carrying a Car8 mutation. Electrophysiological analysis showed that spontaneous excitatory transmission was largely diminished while paired pulse ratio at parallel fiber-PC synapses was comparable to wild-type, suggesting functional synapses have normal release probability but the number of functional synapses may be lower in mutants. Light microscopic study revealed an abnormal extension of climbing fibers to the distal PC dendrites. At the ultrastructural level, we found numerous PC spines not forming synapses primarily in distal dendrites and occasionally multiple spines contacting a single varicosity. These abnormalities of parallel fiber-PC synapses may underlie the functional defect in excitatory transmission. Thus, Car8 plays a critical role in synaptogenesis and/or maintenance of proper synaptic morphology and function in the cerebellum.

Bidirectional dopaminergic modulation of excitatory synaptic transmission in orexin neurons.

Orexin neurons in the lateral hypothalamus (LH)/perifornical area (PFA) are known to promote food intake as well as provide excitatory influence on the dopaminergic reward pathway. Dopamine (DA), in turn, inhibits the reward pathway and food intake through its action in the LH/PFA. However, the cellular mechanism by which DA modulates orexin neurons remains largely unknown. Therefore, we examined the effect of DA on the excitatory neurotransmission to orexin neurons. Whole-cell patch-clamp recordings were performed using acute rat hypothalamic slices, and orexin neurons were identified by their electrophysiological and immunohistochemical characteristics. Pharmacologically isolated action potential-independent miniature EPSCs (mEPSCs) were monitored. Bath application of DA induced a bidirectional effect on the excitatory synaptic transmission dose dependently. A low dose of DA (1 microM) increased mEPSC frequency, which was blocked by the D1-like receptor antagonist SCH 23390, and mimicked by the D1-like receptor agonist SKF 81297. In contrast, higher doses of DA (10-100 microM) decreased mEPSC frequency, which could be blocked with the D2-like receptor antagonist, sulpiride. Quinpirole, the D2-like receptor agonist, also reduced mEPSC frequency. None of these compounds affected the mEPSCs amplitude, suggesting the locus of action was presynaptic. Furthermore, DA (1 microM) induced an increase in the action potential firing, whereas DA (100 microM) hyperpolarized and ceased the firing of orexin neurons, indicating the effect of DA on excitatory synaptic transmission may influence the activity of the postsynaptic cell. In conclusion, our results suggest that D1- and D2-like receptors have opposing effects on the excitatory presynaptic terminals impinging onto orexin neurons.