GABAA receptor modulation of 5-HT neuronal firing in the median raphe nucleus: Implications for the action of anxiolytics

5-HT neurones in the median raphe nucleus (MRN) are involved in anxiety and the sleep/wake cycle. Here, using in vitro electrophysiology, we examined if the firing of MRN 5-HT neurones is regulated by GABA_A receptors. The GABA_A receptor agonists THIP and muscimol caused concentration dependent inhibition of MRN 5-HT neurones. The GABA_A receptor antagonist bicuculline blocked the responses to THIP and muscimol. Bicuculline alone increased the basal firing activity. Responses to THIP were enhanced by the Z hypnotic zolpidem at concentrations selective for the ₂/₃ subunits of the GABA_A receptor (0.2 and 1 μM) but not at a concentration selective for the ₁ subunit (0.02 μM). Consistent with these functional data, 5-HT neurones have been shown to express the ₃ (but not ₂) subunit. The anxiolytic effects of GABA_A receptor modulators are reportedly mediated by ₃-containing receptors. Hence the MRN 5-HT system may be a target for anxiolytic drugs.     

In vivo evidence that 5-HT(2C) receptors inhibit 5-HT neuronal activity via a GABAergic mechanism

BACKGROUND AND PURPOSE: Recent evidence suggests that 5-HT(2C) receptor activation may inhibit midbrain 5-HT neurones by activating neighbouring GABA neurones. This hypothesis was tested using the putative selective 5-HT(2C) receptor agonist, WAY 161503. EXPERIMENTAL APPROACH: The effect of WAY 161503 on 5-HT cell firing in the dorsal raphe nucleus (DRN) was investigated in anaesthetised rats using single unit extracellular recordings. The effect of WAY 161503 on DRN GABA neurones was investigated using double label immunohistochemical measurements of Fos, glutamate decarboxylase (GAD) and 5-HT(2C) receptors. Finally, drug occupancy at 5-HT(2A) receptors was investigated using rat positron emission tomography and ex vivo binding studies with the 5-HT(2A) receptor radioligand [(11)C]MDL 100907. KEY RESULTS: WAY 161503 caused a dose-related inhibition of 5-HT cell firing which was reversed by the 5-HT(2) receptor antagonist ritanserin and the 5-HT(2C) receptor antagonist SB 242084 but not by the 5-HT(1A) receptor antagonist WAY 100635. SB 242084 pretreatment also prevented the response to WAY 161503. The blocking effects of SB 242084 likely involved 5-HT(2C) receptors because the drug did not demonstrate 5-HT(2A) receptor occupancy in vivo or ex vivo. The inhibition of 5-HT cell firing induced by WAY 161503 was partially reversed by the GABA(A) receptor antagonist picrotoxin. Also, WAY 161503 increased Fos expression in GAD positive DRN neurones and DRN GAD positive neurones expressed 5-HT(2C) receptor immunoreactivity. CONCLUSIONS AND IMPLICATIONS: These findings indicate that WAY 161503 inhibits 5-HT cell firing in the DRN in vivo, and support a mechanism involving 5-HT(2C) receptor-mediated activation of DRN GABA neurones.     

GABAergic modulation of 5-HT7 receptor-mediated effects on 5-HT efflux in the guinea-pig dorsal raphe nucleus

5-HT(7) receptor mRNA and protein are localised in the dorsal raphe nucleus (DRN) on non-serotonergic neurones. The effect of 5-HT(7) receptor antagonism on 5-HT efflux was measured from guinea-pig DRN slices, using the technique of fast cyclic voltammetry. The 5-HT(7) receptor antagonist, SB-269970-A, significantly inhibited 5-HT efflux. The GABA(A) receptor agonist, muscimol, significantly inhibited 5-HT efflux, to a similar degree as SB-269970-A. In contrast, the GABA(A) receptor antagonist, bicuculline, significantly increased 5-HT efflux and attenuated the muscimol-induced inhibition. The muscimol and SB-269970-A effects were not additive and in the presence of bicuculline the SB-269970-A-induced inhibition of 5-HT efflux was attenuated. These data suggest that 5-HT(7) receptor antagonist-induced inhibition of 5-HT efflux occurs indirectly via activation of GABA(A) receptors. That is, 5-HT(7) receptors may be located on GABA interneurones and when activated decrease GABA release and hence decrease the inhibitory tone on 5-HT neurones, increasing 5-HT efflux in the DRN. Therefore, in the presence of GABAergic tone 5-HT(7) receptor antagonists would decrease 5-HT release from the DRN.     

Differential regulation of 5-hydroxytryptamine release by GABAA and GABAB receptors in midbrain raphe nuclei and forebrain of rats.

1. Extracellular 5-hydroxytryptamine (5-HT) was determined in dorsal raphe nucleus (DRN), median raphe nucleus (MRN) and nucleus accumbens by use of microdialysis in unanaesthetized rats. 2. Infusion of the gamma-aminobutyric acid (GABA)A receptor agonist muscimol into DRN and MRN resulted in decreased 5-HT in DRN and MRN, respectively. Muscimol infusion into nucleus accumbens had no effect on 5-HT. 3. Infusion of the GABAA receptor antagonist bicuculline into DRN resulted in increased DRN and nucleus accumbens 5-HT. Bicuculline infusion into MRN had no effect on 5-HT. This suggests that endogenous GABA had a tonic, GABAA receptor-mediated inhibitory effect on 5-HT in DRN, but not in MRN. 4. Infusion of the GABAB receptor agonist baclofen into DRN produced a decrease in DRN 5-HT. Baclofen infusion into nucleus accumbens resulted in decreased nucleus accumbens 5-HT. This suggests that GABAB receptors are present in the area of cell bodies and terminals of 5-hydroxytryptaminergic neurones. 5. Infusion of the GABAB receptor antagonists phaclofen and 2-hydroxysaclofen had no effect on midbrain raphe and forebrain 5-HT. This suggests that GABAB receptors did not contribute to tonic inhibition of 5-HT release. 6. In conclusion, 5-HT release is physiologically regulated by distinct subtypes of GABA receptors in presynaptic and postsynaptic sites.     

5-HT1A receptor-mediated autoinhibition does not function at physiological firing rates: evidence from in vitro electrophysiological studies in the rat dorsal raphe nucleus

5-HT(1A)-mediated autoinhibition of neurones in the dorsal raphe nucleus (DRN) is considered to be the principal inhibitory regulator of 5-HT neuronal activity. The activation of this receptor by endogenous 5-HT was investigated using electrophysiological recordings from the rat DRN in vitro. At a concentration which blocked the inhibitory effect of exogenous 5-HT, the 5-HT(1A) antagonist WAY 100635 did not alter basal firing rate or modulate the excitatory response to the alpha(1)-agonist phenylephrine. Blockade of 5-HT reuptake by a concentration of fluoxetine, which enhanced the inhibitory effect of exogenous 5-HT, lowered phenylephrine-induced basal firing presumably due to potentiation of the effect of endogenous 5-HT. However, this effect was not firing rate dependent and neither the proportional increase nor the time-course of the response to a higher concentration of phenylephrine were altered in the presence of fluoxetine. These data suggest that the inhibitory 5-HT(1A) receptor on raphe neurones is neither tonically activated nor plays any role in modulating the response to excitatory transmitters. Thus, at physiological firing rates this receptor does not appear to function as an autoreceptor of serotonergic neurones of the DRN.     

Further pharmacological characterization of 5-HT2C receptor agonist-induced inhibition of 5-HT neuronal activity in the dorsal raphe nucleus in vivo

Background and purpose:

Recent experiments using non-selective 5-hydroxytryptamine (5-HT)_2C receptor agonists including WAY 161503 suggested that midbrain 5-HT neurones are under the inhibitory control of 5-HT_2C receptors, acting via neighbouring gamma-aminobutyric acid (GABA) neurones. The present study extended this pharmacological characterization by comparing the actions of WAY 161503 with the 5-HT_2C receptor agonists, Ro 60-0275 and 1-(3-chlorophenyl) piperazine (mCPP), as well as the non-selective 5-HT agonist lysergic acid diethylamide (LSD) and the 5-HT releasing agent 3,4-methylenedioxymethamphetamine (MDMA).

Experimental approach:

5-HT neuronal activity was measured in the dorsal raphe nucleus (DRN) using extracellular recordings in anaesthetized rats. The activity of DRN GABA neurones was assessed using double-label immunohistochemical measurements of Fos and glutamate decarboxylase (GAD).

Key results:

Ro 60-0175, like WAY 161503, inhibited 5-HT neurone firing, and the 5-HT_2C antagonist SB 242084 reversed this effect. mCPP also inhibited 5-HT neurone firing (∼60% neurones) in a SB 242084-reversible manner. LSD inhibited 5-HT neurone firing; however, this effect was not altered by either SB 242084 or the 5-HT_2A/C receptor antagonist ritanserin but was reversed by the 5-HT_1A receptor antagonist WAY 100635. Similarly, MDMA inhibited 5-HT neurone firing in a manner reversible by WAY 100635, but not SB 242084 or ritanserin. Finally, both Ro 60-0275 and mCPP, like WAY 161503, increased Fos expression in GAD-positive DRN neurones.

Conclusions and implications:

These data strengthen the hypothesis that midbrain 5-HT neurones are under the inhibitory control of 5-HT_2C receptors, and suggest that the 5-HT_2C agonists Ro 60-0175, mCPP and WAY 161503, but not LSD or MDMA, are useful probes of the mechanism(s) involved.     

Role of the medial prefrontal cortex in 5-HT1A receptor-induced inhibition of 5-HT neuronal activity in the rat

1. We examined the involvement of the frontal cortex in the 5-HT2A receptor-induced inhibition of 5-HT neurones in the dorsal raphe nucleus (DRN) of the anaesthetized rat using single-unit recordings complemented by Fos-immunocytochemistry. 2. Both transection of the frontal cortex as well as ablation of the medial region of the prefrontal cortex (mPFC) significantly attenuated the inhibition of 5-HT neurones induced by systemic administration of the 5-HT1A receptor agonist, 8-OH-DPAT (0.5-16 microg kg(-1), i.v.). In comparison, the response to 8-OH-DPAT was not altered by ablation of the parietal cortex. The inhibitory effect of 8-OH-DPAT was reversed by the 5-HT1A receptor antagonist, WAY 100635 (0.1 mg kg(-1), i.v.) in all neurones tested. 3. In contrast, cortical transection did not alter the sensitivity of 5-HT neurones to iontophoretic application of 8-OH-DPAT into the DRN. Similarly, cortical transection did not alter the sensitivity of 5-HT neurones to systemic administration of the selective 5-HT reuptake inhibitor, paroxetine (0.1-0.8 mg kg(-1) , i.v.). 4. 8-OH-DPAT evoked excitation of mPFC neurones at doses (0.5-32 microg kg(-1), i.v.) in the range of those which inhibited 5-HT cell firing. At higher doses (32-512 microg kg(-1), i.v.) 8-OH-DPAT inhibited mPFC neurones. 8-OH-DPAT (0.1 mg kg(-1), s.c.) also induced Fos expression in the mPFC. The neuronal excitation and inhibition, as well as the Fos expression, were antagonized by WAY 100635. 5. These data add further support to the view that the inhibitory effect of 5-HT1A receptor agonists on the firing activity of DRN 5-HT neurones involves, in part, activation of a 5-HT1A receptor-mediated postsynaptic feedback loop centred on the mPFC.     

Effects of idazoxan on dorsal raphe 5-hydroxytryptamine neuronal function.

The effects of the alpha 2-adrenoceptor antagonist idazoxan on 5-hydroxytryptamine (5-HT) neuronal firing and release have been investigated. Idazoxan, administered i.v. (10 micrograms/kg and 0.5 mg/kg) increased dorsal raphe nucleus (DRN)-5-HT neuronal firing rate in a dose-dependent fashion. At the higher dose, a voltammetric study revealed increases in extracellular 5-HT and 5-hydroxyindole acetic acid (5-HIAA) levels, there was no effect with the lower dose. Intra-raphe administration of idazoxan (1 ng) also elevated the firing rate of 5-HT neurones in the dorsal raphe, suggesting that idazoxan may produce the increase in firing by a direct effect in the DRN. However, microiontophoretic application of idazoxan did not increase the firing rate of 5-HT neurones in the DRN. Thus the increase in the firing rate of 5-HT neurones in the DRN observed with systemic and local administration of idazoxan is probably not due to a direct action of idazoxan on the 5-HT neurone. Possibly the idazoxan acted at alpha 2-adrenoceptors located on noradrenergic terminals thus stimulating noradrenaline release and consequently increased 5-HT activity. Chronic administration of idazoxan (0.8 mg/kg per h for 14 days), using osmotic mini-pumps, caused an elevation in basal firing rate and an attenuation of the inhibitory response of DRN 5-HT neurones to the 5-HT1A agonist, 8-hydroxy-2-(di-n-propylamino) tetralin (8-OHDPAT) (10 micrograms/kg i.v.). This finding suggests that chronic infusion with idazoxan leads to desensitisation of the 5-HT1A somatodendritic autoreceptor.     

Inhibition of median and dorsal raphe neurones following administration of the selective serotonin reuptake inhibitor paroxetine

Acute systemic injection of selective serotonin reuptake inhibitors (SSRIs) decreases 5-HT neuronal firing in the dorsal raphe nucleus (DRN). Recent data, however, question whether these drugs also inhibit the firing of 5-HT neurones in the median raphe nucleus (MRN). Using in vivo extracellular electrophysiological recording techniques in the chloral hydrate anaesthetised rat, we have tested the effect of acute administration of the SSRI, paroxetine, on 5-HT neuronal activity in the MRN and DRN. Presumed 5-HT neurones in the MRN displayed the same electrophysiological characteristics as those in the DRN, the only detectable difference being that MRN neurones showed a significantly (p < 0.001) slower mean ( ± SEM(n)) spontaneous firing rate (MRN, 5.6 ± 0.9 (14) spikes/10 s; DRN, 13.5 ± 1.6 (24) spikes/10 s). Paroxetine caused a dose-related (0.1–0.8 mg/kg i.v.) inhibition of all MRN neurones tested (n = 8), producing a complete cessation of cell-firing at the highest doses. DRN neurones (n = 9) responded in a similar fashion. Furthermore, paroxetine inhibited MRN and DRN neurones with almost identical potency (MRN ED₅₀ 259 ± 57 g/kg i.v.: DRN ED₅₀ 243 ± 49 g/kg i.v.). In the majority of cells tested, the effect of paroxetine was reversed by the 5-HT_1A receptor antagonists spiperone or (+)WAY100135, implicating the involvement of the 5-HT_1A autoreceptor. The selective 5-HT_1A receptor agonist 8-OH-DPAT also inhibited the firing of MRN (n = 5) and DRN (n = 12) neurones and with equal potency (MRN ED₅₀, 1.32 ± 0.40 g/kg i.v.: DRN ED₅₀, 1.19 ± 0.23 g/kg i.v.). Our data indicate that paroxetine not only inhibits the firing of 5-HT neurones in the MRN but does so with equal potency to those in the DRN.     

Behavioural and microdialysis study after neurotoxic lesion of the dorsal raphe nucleus in rats

The study investigated the effects of a 5,7-dihydroxytryptamine (5,7-DHT) lesion of the dorsal raphe nucleus (DRN) on anxiety-related behaviour and neurochemical correlates in rats. Behaviour was assessed in the elevated plus maze test (X-maze). Lesion of the DRN reduced markedly 5-HT levels in projection areas by at least 60%. Destruction of the serotonergic neurons in the DRN changed neither anxiety-related behaviour on the elevated plus maze, nor aversion-induced 5-HT release in the brain. Exposure of the lesioned rats to the elevated plus maze increased extracellular 5-HT (148%) in the ventral hippocampus similar as in sham-lesioned (162%) and non-lesioned (160%) controls. The results demonstrate that lesioning of 5-HT neurons in the DRN does not abolish totally the control of anxiety-related behaviour.     

The progesterone metabolite allopregnanolone potentiates GABA(A) receptor-mediated inhibition of 5-HT neuronal activity.

The dorsal raphe nucleus (DRN) is the origin of much of the 5-HT innervation of the forebrain. The activity of DRN 5-HT neurons is regulated by a number of receptors including GABA(A) and 5-HT(1A) inhibitory receptors and by excitatory alpha(1)-adrenoceptors. Using in vitro electrophysiological recording we investigated the action of progesterone and its metabolite, allopregnanolone on receptor-mediated responses of DRN 5-HT neurons. Neither allopregnanolone nor progesterone affected the alpha(1)-adrenoceptor agonist-induced firing. Allopregnanolone also had no effect on the inhibitory response to 5-HT. However, allopregnanolone significantly potentiated the inhibitory responses to GABA(A) receptor agonists. Progesterone did not enhance GABA(A) receptor-meditated inhibitory responses. Thus, the neuroactive metabolite of progesterone, allopregnanolone, has the ability to cause potentiation of GABA(A)-mediated inhibition of DRN 5-HT neurons. This effect on 5-HT neurotransmission may have relevance for mood disorders commonly associated with reproductive hormone events, such as premenstrual dysphoric disorder and postpartum depression.     

Influence of AMPA/kainate receptors on extracellular 5-hydroxytryptamine in rat midbrain raphe and forebrain

1. The regulation of 5-hydroxytryptamine (5-HT) release by excitatory amino acid (EAA) receptors was examined by use of microdialysis in the CNS of freely behaving rats. Extracellular 5-HT was measured in the dorsal raphe nucleus (DRN), median raphe nucleus (MRN), nucleus accumbens, hypothalamus, frontal cortex, dorsal and ventral hippocampus.
2. Local infusion of kainate produced increases in extracellular 5-HT in the DRN and MRN. Kainate infusion into forebrain sites had a less potent effect.
3. In further studies of the DRN and nucleus accumbens, kainate-induced increases in extracellular 5-HT were blocked by the EAA receptor antagonists, kynurenate and 6,7-dinitroquinoxaline-2,3-dione (DNQX).
4. The effect of infusing kainate into the DRN or nucleus accumbens was attenuated or abolished by tetrodotoxin (TTX), suggesting that the increase in extracellular 5-HT is dependent on 5-HT neuronal activity. In contrast, ibotenate-induced lesion of intrinsic neurones did not attenuate the effect of infusing kainate into the nucleus accumbens. Thus, the effect of kainate in the nucleus accumbens does not depend on intrinsic neurones.
5. Infusion of α-amino-3-hydroxy-5-methyl-4-isoxazolaproprionate (AMPA) into the DRN and nucleus accumbens induced nonsignificant changes in extracellular 5-HT. Cyclothiazide and diazoxide, which attenuate receptor desensitization, greatly enhanced the effect of AMPA on 5-HT in the DRN, but not in the nucleus accumbens.
6. In conclusion, AMPA/kainate receptors regulate 5-HT in the raphe and in forebrain sites.

     

Effects of repeated treatment with 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) on the autoregulatory control of dorsal raphe 5-HT neuronal firing and cortical 5-HT release.

These experiments were designed to examine the effects of repeated 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) treatment on the autoregulatory control of cortical 5-HT release and dorsal raphe nucleus (DRN) 5-HT neuronal cell firing. Repeated DOI treatment decreased the behavioural responsiveness (wet-dog shakes) of 5-HT2 receptors and attenuated the inhibitory effects of the 5-HT1A receptor agonist, 8-OH-DPAT (8-hydroxy-2-(di-n-propylamino)tetralin), on both cortical 5-HT release and DRN 5-HT neuronal firing. In contrast, the inhibitory effect of acute DOI on cortical 5-HT release and DRN 5-HT neuronal firing was unaffected by repeated DOI treatment. The results demonstrate that changes in the responsiveness of 5-HT2 receptor function may influence the responsiveness of presynaptic 5-HT1A receptors regulating 5-HT neuronal function. The results also provide further evidence that the inhibition of cortical 5-HT release and DRN 5-HT neuronal firing produced by DOI is not mediated by 5-HT2 receptor activation.     

Evaluation of the anxiolytic-like profile of the GABAB receptor positive modulator CGP7930 in rodents

There is a growing body of data to support the notion that GABA(B) receptors may be a therapeutic target for anxiety disorders. However, the application of GABA(B) receptor agonists in anxiety research and psychiatry is hampered by side effects that include motor in-coordination and hypothermia. Recently the GABA(B) receptor positive modulator GS39783 was shown to be anxiolytic in rodent models, but was devoid of accompanying side effects characteristic of full agonists. However, it is important to test whether such anxiolytic effects generalise to another chemical class of GABA(B) receptor positive modulators. We therefore aimed to investigate the anxiolytic and side-effect profile of CGP7930, the first-reported GABA(B) receptor positive modulator, in rodent models of anxiety, motor coordination and hypothermia. CGP7930 (3-300 mg/kg) showed a modest, compared to the benzodiazepine chlordiazepoxide (10mg/kg), dose-dependent anxiolytic profile in the mouse stress-induced hyperthermia (100mg/kg), staircase (100 and 300 mg/kg) and elevated zero maze tests (3-100mg/kg), but did not have any anxiolytic effects in the rat elevated plus maze. Similar to GS39783, CGP7930 also demonstrated a greatly reduced side-effect profile in comparison to the GABA(B) receptor full agonist baclofen in the mouse rotarod and traction wire tests and did not induce hypothermia. Although the effects of CGP7930 were modest, these results represent a second, structurally distinct, class of GABA(B) positive modulators showing anxiolytic activity. As such, these data support the premise that GABA(B) receptor positive modulation represents a novel therapeutic strategy for the development of anxiolytic drugs with a superior side-effect profile. The generation of more potent compounds is now warranted.     

The role of nucleus accumbens shell GABA receptors on ventral tegmental area intracranial self-stimulation and a potential role for the 5-HT(2C) receptor

Brain γ-aminobutyric acid (GABA) and 5-hydroxytryptamine (5-HT)(2C) receptors are implicated in the neuronal regulation of reward- and aversion-related behaviour. Within the mesocorticolimbic pathways of the brain, relationships between GABA containing neurons and 5-HT(2C) receptor activity may be important in this context. The primary aim of this study was to investigate the role of NAc shell GABA receptors on ventral tegmental area intracranial self-stimulation (ICSS) and to examine the systemic effects of GABAergic ligands in this context. The second aim was to investigate the relationship between GABA receptor- and 5-HT(2C) receptor-related ICSS behaviour, using systemic administration of the selective agonist WAY 161503. Locomotor activity was assessed to compare the potential motor effects of drugs; feeding behaviour and intra-NAc injections of amphetamine (1.0 μg/side) were used as positive controls. When administered systemically the GABA(A) receptor agonist muscimol and antagonist picrotoxin did not selectively change ICSS reward thresholds, although the 5-HT(2C) receptor agonist WAY 161503 (1.0 mg/kg) decreased reward measures. Intra-NAc shell administration of muscimol (225 ng/side) and picrotoxin (125 ng/side), respectively, decreased and increased measures of reward. Intra-NAc shell baclofen (0-225 ng/side; GABA(B) receptor agonist) did not affect any ICSS measures although it increased feeding. Combining picrotoxin and WAY 161503 attenuated the effects of each. These results suggest that a 5-HT(2C) and GABA(A) receptor-mediated neuronal relationship in the NAc shell may be relevant for the regulation of brain reward pathways.     

Interaction between a selective 5-HT1A receptor antagonist and an SSRI in vivo: effects on 5-HT cell firing and extracellular 5-HT.

1. The acute inhibitory effect of selective 5-hydroxytryptamine (serotonin) reuptake inhibitors (SSRIs) on 5-HT neuronal activity may offset their ability to increase synaptic 5-HT in the forebrain. 2. Here, we determined the effects of the SSRI, paroxetine, and a novel selective 5-HT1A receptor antagonist, WAY 100635, on 5-HT cell firing in the dorsal raphé nucleus (DRN), and on extracellular 5-HT in both the DRN and the frontal cortex (FCx). Extracellular electrophysiological recording and brain microdialysis were used in parallel experiments, in anaesthetized rats. 3. Paroxetine dose-dependently inhibited the firing of 5-HT neurones in the DRN, with a maximally effective dose of approximately 0.8 mg kg-1, i.v. WAY 100635 (0.1 mg kg-1, i.v.) both reversed the inhibitory effect of paroxetine and, when used as a pretreatment, caused a pronounced shift to the right of the paroxetine dose-response curve. 4. Paroxetine (0.8 mg kg-1, i.v.), doubled extracellular 5-HT in the DRN, but did not alter extracellular 5-HT in the FCx. A higher dose of paroxetine (2.4 mg kg-1, i.v.) did increase extracellular 5-HT in the FCx, but to a lesser extent than in the DRN. Whereas 0.8 mg kg-1, i.v. paroxetine alone had no effect on extracellular 5-HT in the FCx, in rats pretreated with WAY 100635 (0.1 mg kg-1), paroxetine (0.8 mg kg-1, i.v.) markedly increased extracellular 5-HT in the FCx.(ABSTRACT TRUNCATED AT 250 WORDS)     

SDF-1α/CXCL12 enhances GABA and glutamate synaptic activity at serotonin neurons in the rat dorsal raphe nucleus

The serotonin (5-hydroxytryptamine; 5-HT) system has a well-characterized role in depression. Recent reports describe comorbidities of mood-immune disorders, suggesting an immunological component may contribute to the pathogenesis of depression as well. Chemokines, immune proteins which mediate leukocyte trafficking, and their receptors are widely distributed in the brain, mediate neuronal patterning, and modulate various neuropathologies. The purpose of this study was to investigate the neuroanatomical relationship and functional impact of the chemokine stromal cell-derived factor-1α/CXCL12 and its receptor, CXCR4, on the serotonin dorsal raphe nucleus (DRN) system in the rat using anatomical and electrophysiological techniques. Immunohistochemical analysis indicates that over 70% of 5-HT neurons colocalize with CXCL12 and CXCR4. At a subcellular level, CXCL12 localizes throughout the cytoplasm whereas CXCR4 concentrates to the outer membrane and processes of 5-HT neurons. CXCL12 and CXCR4 also colocalize on individual DRN cells. Furthermore, electrophysiological studies demonstrate CXCL12 depolarization of 5-HT neurons indirectly via glutamate synaptic inputs. CXCL12 also enhances the frequency of spontaneous inhibitory and excitatory postsynaptic currents (sIPSC and sEPSC). CXCL12 concentration-dependently increases evoked IPSC amplitude and decreases evoked IPSC paired-pulse ratio selectively in 5-HT neurons, effects blocked by the CXCR4 antagonist AMD3100. These data indicate presynaptic enhancement of GABA and glutamate release at 5-HT DRN neurons by CXCL12. Immunohistochemical analysis further shows CXCR4 localization to DRN GABA neurons, providing an anatomical basis for CXCL12 effects on GABA release. Thus, CXCL12 indirectly modulates 5-HT neurotransmission via GABA and glutamate synaptic afferents. Future therapies targeting CXCL12 and other chemokines may treat serotonin related mood disorders, particularly depression experienced by immune-compromised individuals.     

Anxiolytic-like effects of Gastrodia elata and its phenolic constituents in mice

The purpose of this study was to characterize the putative anxiolytic-like effects of the aqueous extract of the rhizome of Gastrodia elata along with its phenolic constituents, 4-hydroxybenzyl alcohol (HA) and 4-hyroxybenzaldehyde (HD), using an elevated plus maze (EPM) in mice. The mice were administered either the aqueous G. elata extract orally or received an intraperitoneal injection of the phenolic constituents, 1 h before the behavioral evaluation in the EPM. A single treatment of the aqueous G. elata extract significantly increased the percentage of time spent and arm entries into the open arms of the EPM versus the saline controls. Among the phenolic constituents of G. elata, HA and HD significantly increased the percentage of time spent and arm entries into the open arms of the EPM versus saline controls (p<0.05). Moreover, there were no changes in the locomotor activity and myorelaxant effects in any group compared with the saline controls. In addition, the anxiolytic-like effects of G. elata extract were blocked by both WAY 100635 (0.3 mg/kg, i.p.), a 5-HT(1A) receptor antagonist, and flumazenil (10 mg/kg, i.p.), a GABA(A) receptor antagonist. The anxiolytic-like effects of HA were inhibited by WAY 100635 and the effects of HD were antagonized by flumazenil. These results indicate that G. elata is an effective anxiolytic agent, and suggests that the anxiolytic-like effects of G. elata via the serotonergic nervous system depends on HA and those effects of G. elata via the GABAergic nervous system depends on HD.     

Effects of GABAB receptor ligands in animal tests of depression and anxiety

Preclinical evidence strongly implicates GABA(B) receptors in the pathophysiology of several psychiatric disorders including anxiety and depression. In the present study, we investigated the effects of the selective GABA(B) receptor agonists baclofen and SKF 97541, the GABA(B) receptor positive allosteric modulator CGP7930 and the GABA(B) receptor antagonist SCH 50911 in the modified forced swimming test (FST) and in the elevated zero maze test (EZM), i.e. in animal models predictive of antidepressant and antianxiety activities, respectively. The classical antidepressant imipramine and the anxiolytic diazepam were employed as control drugs in the FST and in the EZM, respectively. In the FST, baclofen (0.25 mg/kg), SKF 97541 (0.01-0.05 mg/kg) or CGP 7930 (1-3 mg/kg) and SCH 50911 (1-3 mg/kg) showed antidepressant-like activity, significantly decreasing immobility time; these effects were not related to changes in locomotor activity. The antidepressant effects produced by the GABA(B) receptor ligands were compared with that of imipramine (30 mg/kg). In the EZM, CGP 7930 (1 mg/kg) and SCH 50911 (1-3 mg/kg) produced anxiolytic-like effects, significantly increasing time spent in the open areas of the maze; those effects were comparable with the effects of diazepam (1-2 mg/kg). Our results indicate that differential manipulation of GABA(B) receptors can modify behaviors relevant to depression and anxiety. The GABA(B) receptor positive allosteric modulator CGP 7930 and the antagonist SCH 50911 show both antidepressant and anxiolytic profile, while the GABA(B) receptor agonists (baclofen and SKF 97541) produce effects that are characteristic of antidepressant drugs.     

The dorsal raphé nucleus is a crucial structure mediating nicotine's anxiolytic effects and the development of tolerance and withdrawal responses

RATIONALE: Smokers frequently report that they obtain anxiety-reducing (anxiolytic) effects from smoking, and this may be one factor which contributes to nicotine dependence. OBJECTIVE: The aim of this study was to investigate the role of the dorsal raphé nucleus (DRN) in mediating the acute anxiolytic effect of nicotine, the development of tolerance to this effect and the anxiogenic response observed on withdrawal from chronic nicotine. METHODS: The social interaction test of anxiety was used to investigate the effects of a range of doses of (-)-nicotine (2.5-4000 ng) following DRN infusion, and whether co-administration of the specific 5-HT1A receptor antagonist WAY 100635 could antagonise the anxiolytic action of nicotine. We then examined the effects of intra-DRN nicotine (2.5-7 ng) following six daily injections of subcutaneous (s.c.) (-)-nicotine (0.1 mg/kg). Finally, we examined whether s.c. or intra-DRN (-)-nicotine could antagonise the anxiogenic response seen 72 h after the termination of 7 days of nicotine treatment. RESULTS: Acute nicotine administration into the DRN produced dose-related effects: low doses (2.5-10 ng) induced an anxiolytic effect, intermediate doses were behaviourally silent (100-1000 ng), and an anxiogenic effect was seen following administration of a high dose (4 micrograms). The anxiolytic effect of (-)-nicotine (5 ng) was reversed by co-administration of a behaviourally inactive dose of WAY 100635 (200 ng). Following 6 days of treatment with s.c. 0.1 mg/kg per day (-)-nicotine, tolerance developed to its anxiolytic action in the DRN. Rats withdrawn for 72 h following this chronic treatment showed an anxiogenic response which was reversed by (-)-nicotine injected s.c. (0.1 mg/kg) or into the DRN (5 ng). CONCLUSIONS: The present findings therefore suggest that the DRN plays an important role in mediating the acute effects of nicotine on anxiety, as measured in the social interaction test, and that the anxiolytic effect is mediated by activation of somatodendritic 5-HT1A autoreceptors. The DRN is also concerned with mediating the development of tolerance to nicotine's anxiolytic effects and because there is an anxiogenic response 72 h after withdrawal from chronic nicotine, this suggests that an oppositional, compensatory mechanism is mediating the tolerance.