Anxiolytic-like Effects and Quantitative EEG Profile of Palmitone Induces Responses Like Buspirone Rather Than Diazepam as Clinical Drugs.

Anxiety is a mental disorder with a growing worldwide incidence due to the SARS-CoV-2 virus pandemic. Pharmacological therapy includes drugs such as benzodiazepines (BDZs) or azapirones like buspirone (BUSP) or analogs, which unfortunately produce severe adverse effects or no immediate response, respectively. Medicinal plants or their bioactive metabolites are a shared global alternative to treat anxiety. Palmitone is one active compound isolated from Annona species due to its tranquilizing activity. However, its influence on neural activity and possible mechanism of action are unknown. In this study, an electroencephalographic (EEG) spectral power analysis was used to corroborate its depressant activity in comparison with the anxiolytic-like effects of reference drugs such as diazepam (DZP, 1 mg/kg) and BUSP (4 mg/kg) or 8-OH-DPAT (1 mg/kg), alone or in the presence of the GABAA (picrotoxin, PTX, 1 mg/kg) or serotonin 5-HT1A receptor antagonists (WAY100634, WAY, 1 mg/kg). The anxiolytic-like activity was assayed using the behavioral response of mice employing open-field, hole-board, and plus-maze tests. EEG activity was registered in both the frontal and parietal cortex, performing a 10 min baseline and 30 min recording after the treatments. As a result, anxiety-like behavior was significantly decreased in mice administered with palmitone, DZP, BUSP, or 8-OH-DPAT. The effect of palmitone was equivalent to that produced by 5-HT1A receptor agonists but 50% less effective than DZP. The presence of PTX and WAY prevented the anxiolytic-like response of DZP and 8-OH-DPAT, respectively. Whereas only the antagonist of the 5-HT1A receptor (WAY) inhibited the palmitone effects. Palmitone and BUSP exhibited similar changes in the relative power bands after the spectral power analysis. This response was different to the changes induced by DZP. In conclusion, brain electrical activity was associated with the anxiolytic-like effects of palmitone implying a serotoninergic rather than a GABAergic mechanism of action.

Tonic serotonergic input increases the burst firing mode and diminishes the firing rate of reticular thalamic nucleus neurons through 5-HT1A receptors activation in anesthetized rats.

The reticular thalamic nucleus (RTn) is a thin shell of GABAergic neurons that covers the dorsal thalamus that regulate the global activity of all thalamic nuclei. RTn controls the flow of information between thalamus and cerebral cortex since it receives glutamatergic information from collaterals of thalamo-cortical (TCs) and cortico-thalamic neurons. It also receives aminergic information from several brain stem nuclei, including serotonergic fibers originated in the dorsal raphe nucleus. RTn neurons express serotonergic receptors including the 5-HT1A subtype, however, the role of this receptor in the RTn electrical activity has been scarcely analyzed. In this work, we recorded in vivo the unitary spontaneous electrical activity of RTn neurons in anesthetized rats; our study aimed to obtain information about the effects of 5-HT1A receptors in RTn neurons. Local application of fluoxetine (a serotonin reuptake inhibitor) increases burst firing index accompanied by a decrease in the basal spiking rate. Local application of different doses of serotonin and 8-OH-DPAT (a specific 5-HT1A receptor agonist) causes a similar response to fluoxetine effects. Local 5-HT1A receptors blockade produces opposite effects and suppresses the effect by 8-OH-DPAT. Our findings indicate the presence of a serotonergic tonic discharge in the RTn that increases the burst firing index and simultaneously decreases the basal spiking frequency through 5-HT1A receptors activation.

Activation of 5-HT1A and 5-HT7 receptors enhanced a positively reinforced long-term memory.

Memory is one of the most important capabilities of our mind since it determines our individuality. Memory formation involves different stages: acquisition, consolidation and retrieval. There are many studies about early stages, however little is known about memory retrieval. Retrieval is the use of learned information and represents a big problem in patients with memory deficits where the main issue is that they can learn but cannot remember. Previous findings have demonstrated that 5-hydroxytryptamine (5-HT) is a neurotransmitter involved in memory process. Hence, here we are exploring the role of 5-HT in memory retrieval by using its metabolic precursor l-tryptophan and several ligands at 5-HT1A and 5-HT7 receptors. Experimental protocol consisted of evaluating conditioned responses (%CR) after one week of interruption following autoshaping sessions for memory formation; a decrease of %CR was interpreted as memory decay. Systemic administration of: (1) l-tryptophan (50 and 100 mg/kg), (2) 5-HT1A receptor agonist 8-OH-DPAT (0.031 and 0.062 mg/kg), (3) the selective antagonist 5-HT1A receptor WAY 100635 (0.3 and 0.6 mg/kg), (4) the 5-HT7 receptor agonist, LP 211, in a dose-dependent manner (1, 2.5, 5.0 and 10.0 mg/kg) enhanced memory retrieval. Further, the 5-HT7 receptor antagonist, SB 269970 (10.0 mg/kg), had no effect. Finally, SB 269970 (10.0 mg/kg) significantly blocked memory retrieval enhancement produced by 10.0 mg/kg LP 211, but not that induced by 2.5 mg/kg LP 211.These results, taken together, suggest that activation of 5-HT1A and 5-HT7 receptors enhanced memory retrieval and these receptors may be therapeutic targets to improve long-term memory retrieval.

Differential modulation of active expiration during hypercapnia by the medullary raphe in unanesthetized rats.

Active expiration represents an important mechanism to improve ventilation in conditions of augmented ventilatory demand, such as hypercapnia. While a rostral ventromedullary region, the parafacial respiratory group (pFRG), has been identified as a conditional expiratory oscillator, little is known about how central chemosensitive sites contribute to modulate active expiration under hypercapnia. In this study, we investigated the influence of the medullary raphe in the emergence of phasic expiratory abdominal activity during hypercapnia in unanesthetized adult male rats, in a state-dependent manner. To do so, reverse microdialysis of muscimol (GABAA receptor agonist, 1 mM) or 8-OH-DPAT (5-HT1A agonist, 1 mM) was applied in the MR during sleep and wakefulness periods, both in normocapnic (room air) and hypercapnic conditions (7% CO2). Electromyography (EMG) of diaphragm and abdominal muscles was performed to measure inspiratory and expiratory motor outputs. We found that active expiration did not occur in room air exposure during wakefulness or sleep. However, hypercapnia did recruit active expiration, and differential effects were observed with the drug dialyses in the medullary raphe. Muscimol increased the diaphragm inspiratory motor output and also increased the amplitude and frequency of abdominal expiratory rhythmic activity during hypercapnia in wakefulness periods. On the other hand, the microdialysis of 8-OH-DPAT attenuated hypercapnia-induced active expiration in a state-dependent manner. Our data suggest that the medullary raphe can either inhibit or potentiate respiratory motor activity during hypercapnia, and the balance of these inhibitory or excitatory outputs may determine the expression of active expiration.

Administration of low doses of the 5-HT1A receptor agonist 8-OH-DPAT attenuates the discriminative signal of amphetamine in the conditioned taste aversion procedure.

Several studies have reported that low doses of the 5-HT1A receptor agonist 8-OH-DPAT reduce cocaine-induced locomotor activity. However, it has also been reported that high doses of 8-OH-DPAT do not substitute for or alter the discriminative signal of cocaine (COC) or amphetamine (AMPH). This study aimed to evaluate the effects of low and high doses of the 5-HT1A agonist 8-OH-DPAT on the discriminative signal of AMPH using conditioned taste aversion as a drug discrimination procedure. Additionally, to establish a correlation between the behavioral effects in drug discrimination and changes in dopamine (DA) and gamma-aminobutyric acid (GABA) concentrations, we evaluated the effect of systemic administration of low or high doses of the 5-HT1A receptor agonist 8-OH-DPAT and of the 5-HT1A receptor antagonist WAY100135 on DA and GABA extracellular concentrations in the nucleus accumbens (nAcc) and ventral tegmental area (VTA), respectively, using cerebral microdialysis. The behavioral results showed that low but not high doses of 8-OH-DPAT produced a reduction in the AMPH-induced discriminative signal, while WAY100135 administration prevented such effects. The microdialysis results showed that a low dose of 8-OH-DPAT decreased extracellular DA concentrations in the nAcc and increased GABA concentrations in the VTA. Pretreatment with WAY100135 prevented these effects. These data support the hypothesis that 5-HT1A receptors modulate the behavioral effects of psychostimulant drugs, such as AMPH, through somatodendritic 5-HT1A autoreceptors in the raphe nucleus indicating that 5-HT1A receptors may be an important target for the development of pharmacological treatments for psychostimulant addiction.

Effects of combined 5-HT2A and cannabinoid receptor modulation on a schizophrenia-related prepulse inhibition deficit in mice.

RATIONALE: Prepulse inhibition of the startle reflex (PPI) is disrupted in several psychiatric disorders including schizophrenia. Understanding PPI pharmacology may help elucidate the pathophysiology of these disorders and lead to better treatments. Given the advantages of multi-target approaches for complex mental illnesses treatment, we have investigated the interaction between receptors known to modulate PPI (5-HT1A and 5-HT2A) and the neuromodulatory endocannabinoid system. OBJECTIVES: To investigate serotonin and cannabinoid receptor (CBR) co-modulation in a model of PPI disruption relevant to schizophrenia METHODS: Male Swiss mice were pretreated with WIN 55,212-2 (CBR agonist), rimonabant (CB1R inverse agonist), 8-OH-DPAT (5-HT1A/7 agonist), and volinanserin (5-HT2A antagonist) or with a combination of a cannabinoid and a serotonergic drug. PPI disruption was induced by acute administration of MK-801. RESULTS: WIN 55,212-2 and rimonabant did not change PPI nor block MK-801-induced deficits. 8-OH-DPAT increased PPI in control mice and, in a higher dose, inhibited MK-801-induced impairments. Volinanserin also increased PPI in control and MK-801-treated mice, presenting an inverted U-shaped dose-response curve. Co-administration of either cannabinoid ligand with 8-OH-DPAT did not change PPI; however, the combination of volinanserin with rimonabant increased PPI in both control and MK-801-exposed mice. CONCLUSIONS: WIN 55,212-2 and rimonabant had similar effects in PPI. Moreover, serotonin and cannabinoid receptors interact to modulate PPI. While co-modulation of CBR and 5-HT1A receptors did not change PPI, a beneficial effect of 5-HT2A and CB1R antagonist combination was detected, possibly mediated through potentiation of 5-HT2A blockade effects by concomitant CB1R blockade.

Serotonin 2C receptors in the basolateral amygdala mediate the anxiogenic effect caused by serotonergic activation of the dorsal raphe dorsomedial subnucleus.

BACKGROUND: Stimulation of serotonergic neurons within the dorsal raphe dorsomedial subnucleus facilitates inhibitory avoidance acquisition in the elevated T-maze. It has been hypothesized that such anxiogenic effect is due to serotonin release in the basolateral nucleus of the amygdala, where facilitation of serotonin 2C receptor-mediated neurotransmission increases anxiety. Besides the dorsal raphe dorsomedial subnucleus, the dorsal raphe caudal subnucleus is recruited by anxiogenic stimulus/situations. However, the behavioral consequences of pharmacological manipulation of this subnucleus are still unknown. AIMS: Investigate whether blockade of serotonin 2C receptors in the basolateral nucleus of the amygdala counteracts the anxiogenic effect caused by the stimulation of dorsal raphe dorsomedial subnucleus serotonergic neurons. Evaluate the effects caused by the excitatory amino acid kainic acid or serotonin 1A receptor-modulating drugs in the dorsal raphe caudal subnucleus. METHODS: Male Wistar rats were tested in the elevated T-maze and light-dark transition tests after intra-basolateral nucleus of the amygdala injection of the serotonin 2C receptor antagonist SB-242084 (6-chloro-2,3-dihydro-5-methyl-N-[6-[(2-methyl-3-pyridinyl)oxy]-3-pyridinyl]-1H-indole-1-carboxyamide dihydrochloride) followed by intra-dorsal raphe dorsomedial subnucleus administration of the serotonin 1A receptor antagonist WAY-100635 (N-[2-[4-2-methoxyphenyl)-1-piperazinyl]ethyl]-N-2-pyridinil-cyclohexanecarboxamide maleate). In the dorsal raphe caudal subnucleus, animals were injected with kainic acid, WAY-100635 or the serotonin 1A receptor agonist 8-OH-DPAT ((±)-8-hydroxy-2-(di-n-propylamino) tetralin hydrobromide) and tested in the elevated T-maze. RESULTS: SB-242084 in the basolateral nucleus of the amygdala blocked the anxiogenic effect caused by the injection of WAY-100635 in the dorsal raphe dorsomedial subnucleus. Kainic acid in the dorsal raphe caudal subnucleus increased anxiety, but also impaired escape expression in the elevated T-maze. Neither WAY-100635 nor 8-OH-DPAT in the dorsal raphe caudal subnucleus affected rat's behavior in the elevated T-maze. CONCLUSION: Serotonin 2C receptors in the basolateral nucleus of the amygdala mediate the anxiogenic effect caused by the stimulation of serotonergic neurons in the dorsal raphe dorsomedial subnucleus. The dorsal raphe caudal subnucleus regulates anxiety- and panic-like behaviors, presumably by a serotonin 1A receptor-independent mechanism.

Facilitation of fear extinction by novelty is modulated by ß-adrenergic and 5-HT1A serotoninergic receptors in hippocampus.

Extinction is the learned inhibition of retrieval of a previously acquired memory and is a major component of exposure therapy, which has attracted much attention because of the use in the treatment of drug addiction, phobias and particularly fear disorders such as post-traumatic stress disorder (PTSD). Exposure to a novel environment before or after extinction training can enhance the extinction of contextual fear conditioning, however the cellular and molecular substrates are still unclear. Here, we investigated the participation of H2-histaminergic, ß-adrenergic and 5-HT1A-serotonergic receptors of the hippocampus on the enhancement of extinction memory caused by novelty. The infusion into the CA1 region of the serotonin 5-HT1A-receptor agonist, 8-OH-DPAT and the ß-adrenergic blocker, Timolol, after the exposure to the novelty hindered the enhancement of extinction by novelty, while Timolol also hindered the extinction consolidation when infused post-extinction. These impairments were abolished by the coinfusion of 8-OH-DPAT plus the 5-HT1A receptor antagonist, NAN-190 and Timolol plus ß-adrenergic agonist, Isoproterenol. However, Dimaprit and Ranitidine blocked the retrieval of CFC, but did not prevented the extinction learning. Here we elucidated some of the molecular mechanisms that are involved on the enhancement of extinction by novelty, demonstrating that the ß-adrenoreceptors and 5-HT1A serotonergic receptors participate on this process alongside with dopaminergic D1 receptors previously described, while histamine H2 receptors, so ubiquitous in learning-related functions in hippocampus are not involved.

Interplay between 5-HT2C and 5-HT1A receptors in the dorsal periaqueductal gray in the modulation of fear-induced antinociception in mice.

The confinement of rodents to the open arm of the elevated-plus maze provokes antinociception (OAA). As a type of defensive reaction, the OAA has been investigated through systemic and intramesencephalic (e.g., dorsal portion of the periaqueductal gray - dPAG) injections of anxiolytic-like drugs [e.g., serotonergic (5-HT) receptor agonists or antagonists]. Here we investigated the effects of (i) intra-dPAG injections of a 5HT2C receptor agonist (MK-212; 0.21 or 0.63 nmol) and antagonist (SB 242084; 0.01, 0.1 or 1.0 nmol); (ii) combined injections of SB 242084 and MK-212 into the dPAG; (iii) combined injections of SB 242084 with 8-OHDPAT (10 nmol) into the dPAG on the OAA in male Swiss mice. Nociception was assessed with the writhing test induced by acetic acid injection. Results showed that (i) intra-dPAG injection of MK-212 (0.63 nmol) increased the OAA; (ii) intra-dPAG SB 242084 (1.0 nmol) prevented the OAA; (iii) SB 242084 (0.1 nmol, a dose devoid of intrinsic effect on nociception) blocked the OAA enhancement provoked by MK-212 and enabled 8-OH-DPAT to prevent the OAA. These results suggest that OAA is mediated by 5-HT2C receptors within the dPAG. Intra-dPAG SB242084 administration provoked similar results on the effects produced by MK-212 and 8-OH-DPAT on OAA. In addition, the dPAG 5-HT1A and 5-HT2C receptors interact each other in the modulation of OAA.

Activation of 5-HT2C (but not 5-HT1A) receptors in the amygdala enhances fear-induced antinociception: Blockade with local 5-HT2C antagonist or systemic fluoxetine.

It is well-known that the exposure of rodents to threatening environments [e.g., the open arm of the elevated-plus maze (EPM)] elicits pain inhibition. Systemic and/or intracerebral [e.g., periaqueductal gray matter, amygdala) injections of antiaversive drugs [e.g., serotonin (5-HT) ligands, selective serotonin reuptake inhibitors (SSRIs)] have been used to change EPM-open arm confinement induced antinociception (OAA). Here, we investigated (i) the role of the 5-HT1A and 5-HT2C receptors located in the amygdaloid complex on OAA as well as (ii) the effects of systemic pretreatment with fluoxetine (an SSRI) on the effects of intra-amygdala injections of 8-OH-DPAT (a 5-HT1A agonist) or MK-212 (a 5-HT2C agonist) on nociception in mice confined to the open arm or enclosed arm of the EPM. Nociception was assessed by the writhing test. Intra-amygdala injections of 8-OH-DPAT (10 nmol) or MK-212 (0.63 nmol) produced a pronociceptive effect and intensified OAA, respectively. Fluoxetine (2.5 mg/kg, intraperitoneally) did not change 8-OH-DPAT effects on nociception but antagonized the enhancement of the OAA produced by MK-212. Interestingly, prior injection of SB 242084 (a selective 5-HT2C antagonist) into the amygdala also blocked the MK-212 effects on OAA. These results indicate that 5-HT may facilitate nociception and intensify OAA, respectively, at 5-HT1A and 5-HT2C receptors located in the amygdala of mice. The impairment produced by systemic fluoxetine on the OAA enhancement provoked by intra-amygdala MK-212 suggests that this type of fear-induced antinociception may be modulated by SSRIs.

Estradiol suppresses ingestive response evoked by activation of 5-HT1A receptors in the lateral hypothalamus of ovariectomized rats.

The present study investigated the effects of estradiol (E2) on ingestive behavior after activation of 5-HT1A receptors in the lateral hypothalamus (LH) of female rats habituated to eat a wet mash diet. Ovariectomized rats treated with corn oil (OVX) or estradiol cypionate (OVX+E) received local injections into the LH of vehicle or an agonist of 5-HT1A receptors, 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT; at a dose of 6 nmol). To determine the involvement of these receptors in food intake, some animals were pretreated with N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl) cyclohexane carboxamide maleate (WAY-100635, a 5-HT1A receptor full antagonist, at a dose of 0.37 nmol), followed by the injection of the agonist 8-OH-DPAT or its vehicle. The results showed that the injection of 8-OH-DPAT into the LH of OVX rats significantly increased food intake, and the duration and frequency of this behavior. The pretreatment with E2 suppressed the hyperphagic response induced by 8-OH-DPAT in OVX animals. The inhibition of 5-HT1A receptors after pretreatment with WAY-100635 blocked the hyperphagic effects evoked by 8-OH-DPAT in OVX. These results indicate that the activity of LH 5-HT1A receptors could be affected by blood E2 levels.

µ-Opioid and 5-HT1A receptors in the dorsomedial hypothalamus interact for the regulation of panic-related defensive responses.

The dorsomedial hypothalamus (DMH) and the dorsal periaqueductal gray (DPAG) have been implicated in the genesis and regulation of panic-related defensive behaviors, such as escape. Previous results point to an interaction between serotonergic and opioidergic systems within the DPAG to inhibit escape, involving µ-opioid and 5-HT1A receptors (5-HT1AR). In the present study we explore this interaction in the DMH, using escape elicited by electrical stimulation of this area as a panic attack index. The obtained results show that intra-DMH administration of the non-selective opioid receptor antagonist naloxone (0.5 nmol) prevented the panicolytic-like effect of a local injection of serotonin (20 nmol). Pretreatment with the selective µ-opioid receptor (MOR) antagonist CTOP (1 nmol) blocked the panicolytic-like effect of the 5-HT1AR agonist 8-OHDPAT (8 nmol). Intra-DMH injection of the selective MOR agonist DAMGO (0.3 nmol) also inhibited escape behavior, and a previous injection of the 5-HT1AR antagonist WAY-100635 (0.37 nmol) counteracted this panicolytic-like effect. These results offer the first evidence that serotonergic and opioidergic systems work together within the DMH to inhibit panic-like behavior through an interaction between µ-opioid and 5-HT1A receptors, as previously described in the DPAG.

5-HT1A receptors of the prelimbic cortex mediate the hormonal impact on learned fear expression in high-anxious female rats.

Hormones highly influence female behaviors. However, research on this topic has not usually considered the variable hormonal status. The prelimbic cortex (PrL) is commonly engaged in fear learning. Connections from and to this region are known to be critical in regulating anxiety, in which serotonin (5-HT) plays a fundamental role, particularly through changes in 5-HT1A receptors functioning. Also, hormone fluctuations can greatly influence anxiety in humans and anxiety-related behavior in rodents, and this influence involves the functioning of 5-HT brain systems. The present investigation sought to determine whether fluctuations in ovarian hormones relative to the estrous cycle would influence the expression of learned fear in female rats previously selected as low- (LA) or high-anxious (HA). Furthermore, we investigate the role of the 5-HT system of the PrL, particularly the 5-HT1A receptors, as a possible modulator of estrous cycle influence on the expression of learned fear through intra-PrL microinjections of 5-HT itself or the full 5-HT1A agonist 8-OH-DPAT (8-hydroxy-2-(di-n-propylamine)tetralin). Behavioral changes were assessed using the fear-potentiated startle (FPS) procedure. The results showed that fear intensity is associated with hormonal decay, being more accentuated during the estrus phase. This increase in fear levels was found to be negatively correlated with the expression of potentiated startle. In rats prone to anxiety and tested during the proestrus and estrus phases, 5-HT mechanisms of the PrL seem to play a regulatory role in the expression of learned fear. These results were not replicated in the LA rats. Similar but less intense results were found regarding the early and late diestrus. Our data indicate that future studies on this subject need to take into account the dissociation between low- and high-responsive females to understand how hormones affect emotional behavior.

Distinct effect of 5-HT1A and 5-HT2A receptors in the medial nucleus of the amygdala on tonic immobility behavior.

The tonic immobility (TI) response is an innate fear behavior associated with intensely dangerous situations, exhibited by many species of invertebrate and vertebrate animals. In humans, it is possible that TI predicts the severity of posttraumatic stress disorder symptoms. This behavioral response is initiated and sustained by the stimulation of various groups of neurons distributed in the telencephalon, diencephalon and brainstem. Previous research has found the highest Fos-IR in the posteroventral part of the medial nucleus of the amygdala (MEA) during TI behavior; however, the neurotransmission of this amygdaloid region involved in the modulation of this innate fear behavior still needs to be clarified. Considering that a major drug class used for the treatment of psychopathology is based on serotonin (5-HT) neurotransmission, we investigated the effects of serotonergic receptor activation in the MEA on the duration of TI. The results indicate that the activation of the 5HT1A receptors or the blocking of the 5HT2 receptors of the MEA can promote a reduction in fear and/or anxiety, consequently decreasing TI duration in guinea pigs. In contrast, blocking the 5HT1A receptors or activating the 5HT2 receptors in this amygdalar region increased the TI duration, suggesting an increase in fear and/or anxiety. These alterations do not appear to be due to a modification of spontaneous motor activity, which might non-specifically affect TI duration. Thus, these results suggest a distinct role of the 5HT receptors in the MEA in innate fear modulation.

Antidepressant-like effect of cannabidiol injection into the ventral medial prefrontal cortex-Possible involvement of 5-HT1A and CB1 receptors.

RATIONALE: Systemic administration of cannabidiol (CBD), the main non-psychotomimetic constituent of Cannabis sativa, induces antidepressant-like effects. The mechanism of action of CBD is thought to involve the activation of 5-HT1A receptors and the modulation of endocannabinoid levels with subsequent CB1 activation. The brain regions involved in CBD-induced antidepressant-like effects remain unknown. The ventral medial prefrontal cortex (vmPFC), which includes the infralimbic (IL) and prelimbic (PL) subregions, receives dense serotonergic innervation and plays a significant role in stress responses. OBJECTIVE: To test the hypothesis that the administration of CBD into the IL or PL would induce an antidepressant-like effect through 5-HT1A and CB1 activation. METHODS: Rats received intra-IL or -PL microinjections of CBD (10-60 nmol/side), 8-OH-DPAT (5-HT1A agonist, 5-10 nmol/side), anandamide (AEA, 0.5 pmol/side) or vehicle (0.2 µl/side) and were submitted to the forced swimming (FST) or to the open field (OFT) tests. Independent CBD-treated groups were pre-treated with WAY100635 (10, 30 nmol/side, 5-HT1A antagonist) or AM251 (10 pmol/side, CB1 antagonist) and submitted to the same tests. An additional group was treated with WAY100635 followed by anandamide. RESULTS: CBD (PL: 10-60 nmol; IL:45-60 nmol) and 8-OH-DPAT (10 nmol) administration significantly reduced the immobility time in the FST, without changing locomotor activity in the OFT. WAY100635 (30 nmol) did not induce effect per se but blocked CBD, 8-OH-DPAT and AEA effects. Additionally, AM251 blocked CBD-effects. CONCLUSION: administration of CBD into the vmPFC induces antidepressant-like effects possibly through indirect activation of CB1 and 5-HT1A receptors.

Medial prefrontal cortex serotonergic and GABAergic mechanisms modulate the expression of contextual fear: intratelencephalic pathways and differential involvement of cortical subregions.

Several lines of evidence indicate that the dorsal hippocampus (dH) and medial prefrontal cortex (mPFC) regulate contextual fear conditioning. The prelimbic (PrL), infralimbic (IL) and the anterior cingulate cortex (ACC) subregions of the mPFC likely play distinct roles in the expression of fear. Moreover, studies have highlighted the role of serotonin (5-hydroxytryptamine, 5-HT)- and γ-aminobutyric acid (GABA)-mediated mechanisms in the modulation of innate fear in the mPFC. The present study characterized dH-mPFC pathways and investigated the role of serotonergic and GABAergic mechanisms of the PrL, IL and ACC-area 1 (Cg1) in the elaboration of contextual fear conditioning using fear-potentiated startle (FPS) and freezing behavior in Rattus norvegicus. The results of neurotracing with microinjections of biotinylated dextran amine into the dH revealed a neural link of the dH with the PrL and ACC. Intra-PrL injections of the 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) and the GABAA receptor-selective agonist muscimol reduced contextual FPS and freezing responses. Intra-Cg1 injections of muscimol but not 8-OH-DPAT decreased FPS and freezing responses. However, neither intra-IL injections of a 5-HT1A agonist nor of a GABAA agonist affected these defensive responses. Labeled neuronal fibers from the dH reached the superficial layers of the PrL cortex and spread to the inner layers of PrL and Cg1 cortices, supporting the pharmacological findings. The present results confirmed the involvement of PrL and Cg1 in the expression of FPS and freezing responses to aversive conditions. In addition, PrL serotoninergic mechanisms play a key role in contextual fear conditioning. This study suggests that PrL, IL and Cg1 distinctively contribute to the modulation of contextual fear conditioning.

Interaction between µ-opioid and 5-HT1A receptors in the regulation of panic-related defensive responses in the rat dorsal periaqueductal grey.

A wealth of evidence indicates that the activation of 5-HT1A and 5-HT2A receptors in the dorsal periaqueductal grey matter (dPAG) inhibits escape, a panic-related defensive behaviour. Results that were previously obtained with the elevated T-maze test of anxiety/panic suggest that 5-HT1A and µ-opioid receptors in this midbrain area work together to regulate this response. To investigate the generality of this finding, we assessed whether the same cooperative mechanism is engaged when escape is evoked by a different aversive stimulus electrical stimulation of the dPAG. Administration of the µ-receptor blocker CTOP into the dPAG did not change the escape threshold, but microinjection of the µ-receptor agonist DAMGO (0.3 and 0.5 nmol) or the 5-HT1A receptor agonist 8-OHDPAT (1.6 nmol) increased this index, indicating a panicolytic-like effect. Pretreatment with CTOP antagonised the anti-escape effect of 8-OHDPAT. Additionally, combined administration of subeffective doses of DAMGO and 8-OHDPAT increased the escape threshold, indicating drug synergism. Therefore, regardless of the aversive nature of the stimulus, µ-opioid and 5-HT1A receptors cooperatively act to regulate escape behaviour. A better comprehension of this mechanism might allow for new therapeutic strategies for panic disorder.

Dorsomedial hypothalamus serotonin 1A receptors mediate a panic-related response in the elevated T-maze.

The dorsomedial hypothalamus (DMH) has long been associated with the regulation of escape, a panic-related defensive response. Previous evidence has shown that the activation of serotonin (5-HT) 1A and 2A receptors impairs escape behavior induced by the electrical stimulation of the same region. In this study we further explore the relationship of the DMH with defense by investigating the effects of 5-HT1A activation on escape behavior generated in male Wistar rats by an ethologically based aversive stimuli, exposure to one of the open arms of the elevated T-maze (ETM). Aside from escape, the ETM also allows the measurement of inhibitory avoidance, a defensive response associated with generalized anxiety disorder. To evaluate locomotor activity, after ETM measurements animals were submitted to an open field. Results showed that intra-DMH administration of the 5-HT1A receptor agonist 8-OH-DPAT inhibited escape expression. Local administration of the 5-HT1A antagonist WAY-100635 by its own was ineffective, but blocked the panicolytic-like effect of 8-OH-DPAT. Chronic (21 days) systemic treatment with imipramine potentiated the anti-escape effect of 8-OH-DPAT. No significant effects of treatment with 8-OH-DPAT or imipramine on avoidance latencies or the number of lines crossed in the open field were found. These results indicate that 5-HT1A receptors within the DMH may play a phasic inhibitory role on ETM escape expression. As previously proposed, facilitation of 5-HT1A-mediated neurotransmission in the DMH may be involved in the mechanism of action of anti-panic compounds.

Serotonergic 5-HT2A/2C receptors are involved in prolactin secretion in hyperestrogenic rats.

Serotonin (5-HT) has been shown to participate in prolactin secretion through a complex process resulting in both positive and negative effects. Estrogen has also been recognized as being involved in this serotonergic effect on prolactin release. Therefore, the aim of the present study was to assess whether estradiol modulates serotonergic involvement in prolactin secretion though 5-HT1A and/or 5-HT2A/2C receptors. Ovariectomized female Wistar rats, treated for 2 weeks with estrogen to induce a hyperprolactinemic status (hyperestrogenic rats) or with sunflower oil vehicle (hypoestrogenic rats), were injected daily with normal saline solution or 6-chloro-2-(1-piperazinyl)pyrazine (MK-212), an 5-HT2A/2C receptor agonist, for 4 days. Other groups of ovariectomized animals received 8-hydroxy-2-(di-N-propylamino)tetralin (8-OH-DPAT) or pindolol, an agonist and antagonist of the 5-HT1A receptor respectively, on the last day of treatment with estrogen or vehicle. Prolactin levels were compared among groups in each experiment under the distinct drug treatments. MK-212 was found to increase prolactin concentrations both in hyper- and hypoestrogenic females compared to controls (p<0.05). In contrast, prolactin levels remained similar to those of controls both in hyperestrogenic animals treated with 8-OH-DPAT and pindolol and in hypoestrogenic rats administered the same treatments. In conclusion, our findings indicate the involvement of 5-HT2A/2C receptors on prolactin release through serotonergic pathways in female animals, especially in hyperestrogenic states.

Conditioned fear in low- and high-anxious rats is differentially regulated by cortical subcortical and midbrain 5-HT(1A) receptors.

Interactions between the prelimbic cortex and the basolateral amygdala underlie fear memory processing, mostly through acquiring and consolidating the learning of a conditioned fear. More recently, studies highlighted the role of the dorsal periaqueductal gray (DPAG) in the modulation of learning fear responses. In addition, extensive data in the literature have signaled the importance of serotonin (5-HT) on fear and anxiety. In the present study, the role of 5-HT neurotransmission of the prelimbic cortex, basolateral amygdala or the DPAG on the unconditioned and conditioned fear responses in rats previously selected as low- (LA) or high-anxious (HA) were assessed through local infusions of 5-HT itself (10nmol/0.2µl) or the selective 5-HT1A agonist 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT - 0.3µg/0.2µl). Behavioral analysis was conducted using the fear-potentiated startle (FPS) procedure. Dependent variables recorded were the latency and amplitude of the unconditioned startle response and FPS. Our findings suggest that, on the prelimbic cortex, 5-HT modulates the expression of conditioned fear response in HA rats and this modulation is dependent on 5-HT1A receptors. This is not true, however, for the basolateral amygdala or the DPAG. In these regions LA but not HA rats were susceptible to the anxiolytic-like effect of 5-HT1A receptor activation. It is thought that the expression of conditioned fear in HA subjects may be dependent on other 5-HT receptors, as the 5-HT1B subtype, and/or changes in other systems such as the GABA and glutamate neurotransmitters. These results increase our understanding of the rostrocaudal influence of 5-HT on the unconditioned and conditioned fear responses in LA and HA subjects and, to some extent, are in disagreement with the theoretical current that emphasizes the role of 5-HT on anxiety, mainly at the subcortical and midbrain levels.