Yukie Kawahara

Tonic regulation of the activity of noradrenergic neurons in the locus coeruleus of the conscious rat studied by dual-probe microdialysis

In the present study, receptor specific compounds were infused via a microdialysis probe in the vicinity of the right locus coeruleus (LC). During the infusion of these compounds, the extracellular content of noradrenaline was recorded in the ipsilateral medial prefrontal cortex (mPFC) with a second microdialysis probe. Agonists and antagonists of various subtypes receptors that have been described to be localized on LC cells, were infused near the LC. The receptors investigated were: alpha2-adrenergic, muscarinic, nicotinic, GABAergic (GABAA and GABAB), glutamatergic (NMDA and non-NMDA). The compounds infused were: clonidine (100 microM), idazoxan (50 microM), bicuculline (50 microM), muscimol (50 microM), baclofen (50 microM), CGP52432 (100 microM), NMDA (300 microM), CPP (300 microM), kainate (100 microM), DNQX (500 microM), oxotremorine (100 microM), atropine (10 microM), nicotine (100 microM) and mecamylamine (100 microM). Evidence was provided that GABAA, NMDA, non-NMDA glutamate, and muscarinic cholinergic receptors in the LC played roles in controlling the activity of noradrenaline neurons. The LC noradrenergic neurons were not tonically excitated by glutamatergic or cholinergic afferent neurons, and were not tonically inhibited by alpha2 autoreceptors. Tonic inhibition was evident for GABAergic neurons, acting via GABAA receptors.

The role of afferents to the locus coeruleus in the handling stress-induced increase in the release of noradrenaline in the medial prefrontal cortex : a dual-probe microdialysis study in the rat brain

This study was aimed to identify the neuronal pathways that mediate the handling stress-induced increase in the release of noradrenaline in the medial prefrontal cortex of the rat brain. For that purpose a microdialysis probe was implanted in the vicinity of the locus coeruleus and a second probe was placed in the ipsilateral medial prefrontal cortex. Receptor specific antagonists acting on the alpha(2)-adrenoceptor (50 microM idazoxan), GABA(A) (50 microM bicuculline), GABA(B) (100 microM (3, 4-Dichlorophenyl)methyl]propyl](diethoxymethyl) phosphonic acid; CGP 52432), acetylcholine (10 microM atropine), corticotropin releasing factor (CRF) (100 microM butyl-ethyl-[2,5-dimethyl-7-(2,4, 6-trimethyl-phenyl)-7H-pyrrolo[2,3-d]pyrimidin-4-yl]-amine; CP-154, 526), NMDA glutamate (300 microM (+/-)-3(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid; CPP) and non-NMDA glutamate receptors (500 microM 6,7-dinitroquinoxaline-2, 3-dione; DNQX) were infused into the locus coeruleus by retrograde dialysis, whereas extracellular noradrenaline was recorded in the ipsilateral medial prefrontal cortex. During infusion of the various compounds rats were gently handled for 10 min. Infusion of idazoxan potentiates the handling-induced increase in the release of noradrenaline in the medial prefrontal cortex. The infusions of, atropine, bicuculline, CGP 52432 and DNQX were without effect on the handling response. Infusion of the NMDA receptor antagonist CPP or the non-peptide CRF receptor antagonist CP-154,526 suppressed the stimulation of noradrenaline during stress. It is concluded that alpha(2)-adrenoceptors, NMDA glutamate receptors and CRF receptors modify the handling stress response of locus coeruleus neurones. The data suggest no major role for glutamatergic, GABAergic, or cholinergic afferents to the locus coeruleus in mediating the stress response.

The locus coeruleus noradrenergic system in the rat brain studied by dual-probe microdialysis

A dual-probe microdialysis technique was applied to the locus coeruleus (LC) and prefrontal cortex (PFC) of the brain of conscious rats. One probe was implanted close to the LC and was used to apply receptor-specific compounds by retrograde microdialysis. The effects of the LC infusions were recorded by a sampling noradrenaline by a second probe that was implanted in the ipsilateral prefrontal cortex. Infusion of sodium channel blocker tetrodotoxin (1 microM; 90 min) into the LC decreased extracellular noradrenaline in the PFC to approximately 20% of control values. Infusion of alpha2-adrenoceptor agonist clonidine (100 microM, infused during 15 or 45 min) near to the LC, decreased extracellular noradrenaline in the PFC to 35 and 20% of controls, respectively. These results indicate that > 80% of the extracellular levels of noradrenaline in the PFC is derived from LC innervation, and confirms the importance of alpha2-autoreceptors on noradrenergic neurons in the LC. Infusion of the cholinergic receptor agonist, carbachol (100 microM, 45 min) near to the LC increased extracellular noradrenaline in the PFC to approximately 150% of controls. Infusions of the excitatory amino-acid agonists NMDA and kainate into the LC caused marked increases in extracellular noradrenaline in the PFC to 240 and 200% of controls, respectively. The experiments with clonidine, carbachol, NMDA and kainate were repeated in anesthetized rats. Clonidine and carbachol were similarly effective as in conscious animals but the effects of NMDA and kainate on extracellular noradrenaline in the PFC were clearly suppressed: 145 and 130% of controls, respectively. These results suggest that increased arousal or behavioural activation might have contributed to the increases in extracellular noradrenaline that was seen after infusion of the glutamate agonists. These results also provide evidence for localization of cholinergic-, NMDA-, non-NMDA-receptor on noradrenergic neurons in the LC. Finally it is concluded that dual-probe microdialysis is a useful method to further investigate the pharmacology of LC-noradrenergic neurons. Carbachol and clonidine are suitable tools for a rapid and reversible stimulation or inhibition, respectively, of noradrenergic LC neurons.

The noradrenaline-dopamine interaction in the rat medial prefrontal cortex studied by multi-probe microdialysis

Multi-probe microdialysis was used to investigate the interaction between the release of noradrenaline and dopamine in the medial prefrontal cortex. Retrograde microdialysis was used to stimulate or inhibit the activity of the locus coeruleus for a restricted period of time, and the response of extracellular noradrenaline and dopamine in the ipsilateral and contralateral medial prefrontal cortex was recorded with microdialysis probes. Infusion of clonidine into the locus coeruleus (100 microM for 45 min) suppressed noradrenaline release and slightly inhibited dopamine release in the ipsilateral medial prefrontal cortex. Application of carbachol to the locus coeruleus (100 microM for 45 min) stimulated both the noradrenaline and dopamine release in the ipsilateral medial prefrontal cortex. No changes were seen in the contralateral medial prefrontal cortex. In the ipsilateral nucleus accumbens, extracellular noradrenaline levels increased, but dopamine levels remained unchanged. Application to the locus coeruleus (during 10 min) of the glutamate receptor agonists N-methyl-D-aspartate (NMDA) (300 microM) or kainate (100 microM) strongly increased extracellular noradrenaline and dopamine levels in the ipsilateral medial prefrontal cortex. However, in the contralateral probe the release of dopamine (but not of noradrenaline) was also stimulated. Application of carbachol to the locus coeruleus was used as a model to further investigate the presumed noradrenaline-dopamine interaction. In a series of dual-probe experiments, alpha(1)-, alpha(2)-, and beta-adrenoceptor antagonists (prazosin, idazoxan, propranolol) or a reuptake-inhibitor (nomifensine) was administered during carbachol stimulation of the locus coeruleus. Prazosin and propranolol were administered systemically in a dose of 3 mg/kg, whereas idazoxan (10 microM) and nomifensine (100 microM) were infused into the medial prefrontal cortex. However, none of these pretreatments modified the effects of the control carbachol-infusions. The results did not identify a receptor-interaction or a common reuptake site that explained the presumed interaction between dopamine and noradrenaline in the medial prefrontal cortex. Therefore, the noradrenaline-dopamine interaction hypothesis could not be confirmed or refuted.

Peripherally administered ghrelin induces bimodal effects on the mesolimbic dopamine system depending on food-consumptive states.

Ghrelin induces orexigenic behavior by activation of growth hormone secretagogue 1 receptors (GHSRs) in the ventral tegmental area (VTA) as well as hypothalamus, suggesting the involvement of mesolimbic dopamine system in the action of ghrelin. The present study aimed to identify neuronal mechanisms by which peripherally administered ghrelin regulates the mesolimbic dopamine system under different food-consumptive states. Ghrelin was administered to rats peripherally (3 nmol, i.v.) as well as locally into the VTA (0.3 nmol). Dopamine in the nucleus accumbens shell (NAc) was measured by microdialysis. Peripheral administration of ghrelin decreased dopamine levels in the NAc when food was removed following ghrelin administration. This inhibitory effect was mediated through GABA(A) and N-methyl-D-aspartate (NMDA) receptors in the VTA. In contrast, when animals consumed food following ghrelin administration, dopamine levels increased robustly. This stimulatory effect was mediated through NMDA receptors, but not through GABA(A) receptors, in the VTA. Importantly, both the inhibitory and stimulatory effects of ghrelin primarily required activation of GHSRs in the VTA. Furthermore, local injection of ghrelin into the VTA induced dopamine release in the NAc and food consumption, supporting the local action of ghrelin in the VTA. In conclusion, peripherally administered ghrelin activates GHSRs in the VTA, and induces bimodal effects on mesolimbic dopamine neurotransmission depending on food-consumptive states.

Regional distribution and the dynamics of n-decanoyl ghrelin, another acyl-form of ghrelin, upon fasting in rodents.

n-Decanoyl ghrelin (D-ghrelin), a member of ghrelin-derived peptides, is found in plasma and the stomach; however, there have so far been no studies describing its dynamics. A D-ghrelin-specific radioimmunoassay was established to examine the tissue distribution and the kinetics of D-ghrelin in mice. The effect of D-ghrelin on food intake was also examined and compared to n-octanoyl ghrelin (O-ghrelin). D-ghrelin was detected throughout the gastrointestinal tissue and plasma with highest level in the stomach. An immunofluorescent study revealed the co-localization of D- and O-ghrelin in the same stomach cells. Upon fasting, the levels of D-ghrelin in the stomach and plasma significantly increased, while that of O-ghrelin in the stomach declined. D-ghrelin increased the 2 h food consumption in mice as O-ghrelin does. These findings indicate that D-ghrelin is mainly produced in the stomach to work in concert with O-ghrelin. The different kinetics of D- and O-ghrelin in the stomach upon fasting implies the possibility of D-ghrelin-specific bioregulation.

Upregulation of the dorsal raphe nucleus-prefrontal cortex serotonin system by chronic treatment with escitalopram in hyposerotonergic Wistar-Kyoto rats

Wistar-Kyoto (WKY) rats are sensitive to chronic stressors and exhibit depression-like behavior. Dorsal raphe nucleus (DRN) serotonin (5-HT) neurons projecting to the prefrontal cortex (PFC) comprise the important neurocircuitry underlying the pathophysiology of depression. To evaluate the DRN-PFC 5-HT system in WKY rats, we examined the effects of escitalopram (ESCIT) on the extracellular 5-HT level in comparison with Wistar rats using dual-probe microdialysis. The basal levels of 5-HT in the DRN, but not in the PFC, in WKY rats was reduced as low as 30% of Wistar rats. Responses of 5-HT in the DRN and PFC to ESCIT administered systemically and locally were attenuated in WKY rats. Feedback inhibition of DRN 5-HT release induced by ESCIT into the PFC was also attenuated in WKY rats. Chronic ESCIT induced upregulation of the DRN-PFC 5-HT system in WKY rats, with increases in basal 5-HT in the DRN, responsiveness to ESCIT in the DRN and PFC, and feedback inhibition, whereas downregulation of these effects was induced in Wistar rats. Thus, the WKY rat is an animal model of depression with low activity of the DRN-PFC 5HT system. The finding that chronic ESCIT upregulates the 5-HT system in hyposerotonergic WKY rats may contribute to improved understanding of mechanisms of action of antidepressants, especially in depression with 5-HT deficiency.

The spontaneously hypertensive rat/Izm (SHR/Izm) shows attention deficit/hyperactivity disorder-like behaviors but without impulsive behavior: Therapeutic implications of low-dose methylphenidate

The spontaneously hypertensive rat (SHR) has been used as a genetic animal model of attention deficit/hyperactivity disorder (ADHD). SHR/Izm is derived from stroke-resistant SHR as SHR/NIH and SHR/NCrl but from 22nd to 23rd generation descendants of the SHR/NIH ancestor and therefore may show different behavioral phenotypes compared to other SHR sub-strains. In this study, ADHD-like behaviors in SHR/Izm were evaluated compared to Wistar rats. SHR/Izm showed high locomotor activity in the habituation phase in a novel environment, although locomotor activity in the initial exploratory phase was low. In a behavioral test for attention, spontaneous alternation behavior in the Y-maze test was impaired in SHR/Izm. However, impulsive behavior in the elevated-plus maze test, which is designed to detect anxiety-related behavior but also reflects impulsivity for novelty seeking, was comparable to Wistar rats. Hyperactivity and inattention, detected as ADHD-like behaviors in SHR/Izm, were ameliorated with methylphenidate at a low dose (0.05mg/kg, i.p.). Therefore, SHR/Izm represents a unique animal model of ADHD without anxiety-related impulsive behavior.

Acute effects of resveratrol to enhance cocaine-induced dopamine neurotransmission in the striatum.

Resveratrol is known as an activator of SIRT1, which leads to the deacetylation of histone and non-histone protein substrates, but also has other pharmacological profiles such as the inhibition of monoamine oxidase (MAO)-A and MAO-B. Resveratrol was previously demonstrated to potentiate the rewarding effects of chronic cocaine via activation of SIRT1. However, the role of resveratrol in cocaine responses in the acute phase remains unexplored. Therefore, we investigated the acute effects of resveratrol on cocaine-stimulated dopamine neurotransmission by analyzing protein phosphorylation in neostriatal slices. Treatment with resveratrol (50μM for 30min) enhanced cocaine-induced increases in the phosphorylation of DARPP-32 at Thr34 and GluA1 at Ser845, postsynaptic substrates for dopamine/D1 receptor/PKA signaling, and a cocaine-induced decrease in the phosphorylation of tyrosine hydroxylase at Ser40, a presynaptic substrate for dopamine/D2 receptor signaling. The inhibition of both MAO-A and MAO-B by clorgyline and pargyline, respectively, enhanced the effects of cocaine on DARPP-32 phosphorylation. The acute effect of resveratrol on cocaine-induced DARPP-32 phosphorylation was occluded with inhibition of MAO-A and MAO-B. In behavioral studies, resveratrol (40mg/kg, s.c.) enhanced the increase in locomotor activity induced by acute cocaine administration (10mg/kg, i.p.). Thus, this study provides pharmacological evidence that acute resveratrol enhances cocaine-induced dopamine neurotransmission and behavioral responses, presumably via mechanisms involving the inhibition of dopamine catabolism by MAO-A and MAO-B. Resveratrol may be useful to treat dysregulated dopamine neurotransmission, but it may enhance the risk of developing drug addiction.

Long-term citalopram treatment alters the stress responses of the cortical dopamine and noradrenaline systems: the role of cortical 5-HT1A receptors

Background: Cortical dopamine and noradrenaline are involved in the stress response. Citalopram, a selective serotonin reuptake inhibitor, has direct and indirect effects on the serotonergic system. Furthermore, long-term treatment with citalopram affects the dopamine and noradrenaline systems, which could contribute to the therapeutic action of antidepressants. Methods: The effects of long-term treatment with citalopram on the responses of the dopamine and noradrenaline systems in the rat prefrontal cortex to acute handling stress were evaluated using in vivo microdialysis. Results: Acute handling stress increased dopamine and noradrenaline levels in the prefrontal cortex. The dopamine and noradrenaline responses were suppressed by local infusion of a 5-HT1A receptor agonist, 7-(Dipropylamino)-5,6,7,8-tetrahydronaphthalen-1-ol;hydrobromide, into the prefrontal cortex. The dopamine response was abolished by long-term treatment with citalopram, and the abolished dopamine response was reversed by local infusion of a 5-HT1A receptor antagonist, (Z)-but-2-enedioic acid;N-[2-[4-(2-methoxyphenyl)piperazin-1-yl]ethyl]-N-pyridin-2-ylcyclohexanecarboxamide into the prefrontal cortex. On the other hand, long-term treatment with citalopram reduced the basal noradrenaline levels (approximately 40% of the controls), but not the basal dopamine levels. The noradrenaline response was maintained despite the low basal noradrenaline levels. Signaling from the 5-HT1A receptors and α2-adrenoceptors was not involved in the decrease in the basal noradrenaline levels but partially affected the noradrenaline response. Conclusions: Chronic citalopram treatment differentially suppresses the dopamine and noradrenaline systems in the prefrontal cortex, and the dopamine stress response was preferentially controlled by upregulating 5-HT1A receptor signaling. Our findings provide insight into how antidepressants modulate the dopamine and noradrenaline systems to overcome acute stress.