New evidence for refinement of anesthetic choice in procedures preceding the forced swimming test and the elevated plus-maze.

Previous studies indicated that some general anesthetics induce long-term antidepressant and/or anxiolytic-like effects. This raises the concern about the use of anesthesia in surgeries that precede psychopharmacological tests, since it may be a potential bias on results depending on the experimental design used. Thus, we evaluated whether general anesthetics used in surgeries preceding psychopharmacological tests would affect rats behavior in tests predictive of antidepressant or anxiolytic-like effects. We tested if a single exposure to sub-anesthetic or anesthetic doses of tribromoethanol, chloral hydrate, thiopental or isoflurane would change rats behavior in the forced swimming test (FST) or in the elevated plus-maze (EPM) test, at 2 h or 7 days after their administration. We also evaluated whether prior anesthesia would interfere in the detection of the antidepressant-like effect of imipramine or the anxiolytic-like effect of diazepam. Previous anesthesia with the aforementioned anesthetics did not change rats behaviors in FST per se nor it changed the antidepressant-like effect induced by imipramine treatment. Rats previously anesthetized with tribromoethanol or chloral hydrate exhibited, respectively, anxiogenic-like and anxiolytic-like behaviors in the EPM. Prior anesthesia with thiopental or isoflurane did not produce any per se effect in rats behaviors in the EPM nor disturbed the anxiolytic-like effect of diazepam. Our results suggest that, in our experimental conditions, tribromoethanol and chloral hydrate are improper anesthetics for surgeries that precede behavioral analysis in the EPM. Isoflurane or thiopental may be suitable for anesthesia before evaluation in the EPM or in the FST.

Dorsal hippocampal galanin modulates anxiety-like behaviours in rats.

Galanin, a peptide expressed in mammalian brain regions, has been implicated in anxiety and depression. Galanin signalling occurs through three G protein-linked receptors (GAL1, GAL2 and GAL3). Galanin regulates the release of neurotransmitters in some brain regions related to anxiety, including the hippocampus. GAL2 is the most abundant galanin receptor in the dorsal hippocampus. In this study, we evaluated whether galanin administered in the dorsal hippocampus affected anxiety-like behaviours of rats. We also investigated if GAL2 receptors are involved in the anxiogenic-like effect of galanin using a GAL2 antagonist, M871. To achieve these objectives, male adult Wistar rats received intra-dorsal hippocampal delivery of galanin (0.3 and 1.0 nmol/0.5 µl) or vehicle in experiment 1 and GAL2 antagonist M871 (1.0 and 3.0 nmol/0.5 µl) or vehicle in experiment 2. Twenty min after administration of drugs, the animals were tested in the elevated plus-maze (EPM). Galanin (1.0 nmol) induced anxiogenic-like behaviours, while the GAL2 receptor antagonist M871 (3.0 nmol) induced anxiolytic-like behaviours in rats exposed to the EPM, indicating a tonic effect of galanin. In experiment 3, we evaluated whether previous infusion of the GAL2 antagonist M871 (1 or 2 nmol) in the dorsal hippocampus would block the anxiogenic-like effect of galanin in rats tested in the EPM. We showed that M871 (2.0 nmol) counteracted the anxiogenic-like effect of galanin infused in the dorsal hippocampus of rats. Altogether, our results provide evidence that galanin promotes pharmacological and tonic anxiogenic-like effects in the dorsal hippocampus, possibly mediated by GAL2 receptors.

Galanin subtype 1 and subtype 2 receptors mediate opposite anxiety-like effects in the rat dorsal raphe nucleus.

About 40% of the dorsal raphe nucleus (DRN) neurons co-express serotonin (5-HT) and galanin. Serotonergic pathways from the DRN to the amygdala facilitate learned anxiety, while those from the DRN to the dorsal periaqueductal grey matter (DPAG) impair innate anxiety. Previously, we showed that galanin infusion in the DRN of rats induces anxiolytic effect by impairing inhibitory avoidance without changing escape behaviour in the elevated T-maze (ETM). Here, we evaluated: (1) which galanin receptors would be involved in the anxiolytic effect of galanin in the DRN of rats tested in the ETM; (2) the effects of galanin intra-DRN on panic-like behaviours evoked by electrical stimulation of the DPAG. The activation of DRN GAL1 receptors by M617 (1.0 and 3.0nmol) facilitated inhibitory avoidance, whereas the activation of GAL2 receptors by AR-M1896 (3.0nmol) impaired the inhibitory avoidance in the ETM, suggesting an anxiogenic and an anxiolytic-like effect respectively. Both agonists did not change escape behaviour in the ETM or locomotor activity in the open field. The anxiolytic effect of AR-M1896 was attenuated by the prior administration of WAY100635 (0.18nmol), a 5-HT1A antagonist. Galanin (0.3nmol) administered in the DRN increased discreetly flight behaviours induced by electrical stimulation of the DPAG, suggesting a panicolytic effect. Together, our results showed that galanin mediates opposite anxiety responses in the DRN by activation of GAL1 and GAL2 receptors. The anxiolytic effect induced by activation of Gal2 receptors may depend on serotonergic tone. Finally, the role of galanin in panic related behaviours remains uncertain.

Galanin microinjection into the dorsal periaqueductal gray matter produces paradigm-dependent anxiolytic effects.

Galanin is a peptide that is present in the central nervous system in mammals, including rodents and humans. The actions of galanin are mediated by three types of metabotropic receptors: GAL1, GAL2, and GAL3. GAL1 and GAL3 increase K(+) efflux, and GAL2 increases intracellular Ca(2+) levels. The distribution of galanin and its receptors suggests its involvement in fear and/or anxiety. The periaqueductal gray matter (PAG) is a key mediator of defensive behaviors that is both targeted by galaninergic projections and supplied with GAL1 receptors and, less markedly, GAL2 receptors. We examined the effects of galanin microinjections in the dorsal PAG (dPAG) on the performance of rats in different models of anxiety. Male Wistar rats (n=7-12) were implanted with guide cannulae in the dPAG. They received microinjections of either galanin (0.3, 1.0, and 3.0 nmol) or vehicle and were tested in the Vogel conflict test (VCT), elevated plus maze (EPM), and elevated T-maze (ETM). Rats that were tested in the ETM were further evaluated for exploratory activity in the open field test (OFT). Galanin microinjections had no effects on anxiety-like behavior in the EPM or VCT or exploratory activity in the EPM or OFT. In the ETM, however, microinjections of 3 nmol galanin impaired learned anxiety (i.e., avoidance of the open arms) without changing unconditioned fear (i.e., escape from the open arms). The present data suggest that galanin transmission in the dPAG inhibits the acquisition of anxiety-like responses in the ETM.

Role of nitric oxide in brain regions related to defensive reactions.

Nitric oxide synthase (NOS) positive neurons are located in most brain areas related to defensive reactions, including the dorsolateral periaqueductal grey (dlPAG). NOS inhibitors injected into this structure induce anxiolytic-like responses whereas NO donors promote flight reactions. Intra-dlPAG administration of carboxy-PTIO, a NO scavenger, or ODQ, a soluble guanylate cyclase inhibitor, produced anxiolytic-like effects on rats exposed to the elevated plus-maze (EPM). A double-staining experiment using NADPHd histochemistry and c-Fos immunohistochemistry in rats exposed to a cat or to the EPM showed increased activation of NO producing neurons in the dlPAG, paraventricular and lateral nuclei of hypothalamus and dorsal raphe nucleus. Cat exposure also increased activation of NOS neurons in the medial amygdala, dorsal pre-mammillary nucleus and bed nucleus of stria terminalis. Local infusion into the dlPAG of a glutamate NMDA-receptor antagonist (AP7) or a benzodiazepine agonist (midazolam) completely prevented the flight reactions induced by intra-dlPAG administration of SIN-1, a NO donor. The responses were also inhibited by the 5-HT2A/C agonist DOI but not by a 5-HT1A agonist. These results suggest a modulatory role for NO on brain areas related to defensive reactions, probably by interacting with glutamate, serotonin and/or GABA-mediated neurotransmission.

Effect of Hypericum perforatum on marble-burying by mice.

The effect of Hypericum perforatum extract (LI 160) at a dose that exerts an antidepressive-like effect was studied in mice in the marble-burying test. Acute Hypericum perforatum (150, 300 and 500 mg/kg, p.o.) reduced immobility time in the forced swimming test. The number of marbles buried, but not locomotor activity, was reduced by acute treatment with Hypericum perforatum (150 and 300 mg/kg, p.o.). However, this effect was not seen after chronic treatment (21 days) with Hypericum perforatum (300 mg/kg, p.o.). Thus, Hypericum perforatum extract, at antidepressant dose, exerts an acute anxiolytic drug effect on the marble-burying test, which could indicate a potential anti-obsessive effect, although the development of tolerance could be an important drawback.

The antidepressive-like effect of oxcarbazepine: possible role of dopaminergic neurotransmission.

It has been previously shown that oxcarbazepine (OXCBZ), a keto-analogue of carbamazepine, exhibits an antidepressive-like effect profile in the learned helplessness and forced swimming test (FST). Since carbamazepine possesses dopaminergic effect, the present study was carried out to evaluate the extent to which the antidepressive effect of OXCBZ might be mediated by dopaminergic system. Thus, the effects of OXCBZ in haloperidol-induced catalepsy and apomorphine-induced stereotypy were studied. The anti-immobility effect of OXCBZ in the FST was also evaluated in haloperidol pre-treated rats. OXCBZ (40 and 80 mg/kg, i.p.) dose-dependently reduced the catalepsy induced by haloperidol (2.0 mg/kg, i.p.). Moreover, OXCBZ (80 mg/kg, but not 20 or 40 mg/kg, i.p.) increased the intensity of apomorphine-induced stereotypy (0.6 mg/kg, s.c.). Finally, it was observed that the combination of OXCBZ (80 mg/kg, i. p.) and haloperidol (0.5 mg/kg, i.p.) antagonized the anti-immobility effect of OXCBZ and further increased the immobility time when compared to haloperidol alone. Haloperidol alone (0.5 or 1. 0 mg/kg) did not change the immobility time. Thus, these results suggest that OXCBZ could enhance dopaminergic neurotransmission, which might mediate its antidepressive-like effect.

The effect of oxcarbazepine on behavioural despair and learned helplessness.

The effect of oxcarbazepine was evaluated in two tests of depression (forced swimming and learned helplessness) and in the open-field test. Acute (three times over 24 h) oxcarbazepine 80 mg/kg (but not 40 mg/kg) decreased immobility time in the forced swimming test. In the learned helplessness test, 4 days of treatment with oxcarbazepine 80 mg/kg reversed the deficits induced by foot-shock in rats submitted to the two-way active avoidance test. Oxcarbazepine 80 mg/kg did not modify the behaviour of rats in the open-field test, an indication that, at this dose, oxcarbazepine did not show a locomotor stimulatory effect. Thus, the data of the present study suggest that oxcarbazepine has a potential antidepressive effect.