Neuroprotective Profile of Triazole Grandisin Analogue against Amyloid-Beta Oligomer-Induced Cognitive Impairment.

Alzheimer's disease (AD) is a neurodegenerative disorder caused by accumulation of amyloid-ß oligomers (AßO) in the brain, neuroinflammation, oxidative stress, and cognitive decline. Grandisin, a tetrahydrofuran neolignan, exhibits relevant anti-inflammatory and antioxidant properties. Interestingly, grandisin-based compounds were shown to prevent AßO-induced neuronal death in vitro. However, no study has assessed the effect of these compounds on the AD animal model. This study focuses on a triazole grandisin analogue (TGA) synthesized using simplification and bioisosteric drug design, which resulted in improved potency and solubility compared with the parent compound. This study aimed to investigate the possible in vivo effects of TGA against AßO-induced AD. Male C57/Bl6 mice underwent stereotaxic intracerebroventricular AßO (90 µM) or vehicle injections. 24 h after surgery, animals received intraperitoneal treatment with TGA (1 mg/kg) or vehicle, administered on a 14 day schedule. One day after treatment completion, a novel object recognition task (NORT) was performed. Memantine (10 mg/kg) was administered as a positive control. NORT retention sessions were performed on days 8 and 16 after AßO injection. Immediately after retention sessions, animals were euthanized for cortex and hippocampus collection. Specimens were subjected to oxidative stress and cytokine analyses. TGA reduced the level of cortex/hippocampus lipoperoxidation and prevented cognitive impairment in AßO-injected mice. Additionally, TGA reduced tumor necrosis factor (TNF) and interferon-γ (IFN-γ) levels in the hippocampus. By contrast, memantine failed to prevent cortex/hippocampus lipid peroxidation, recognition memory decline, and AßO-induced increases in TNF and IFN-γ levels in the hippocampus. Thus, memantine was unable to avoid the AßO-induced persistent cognitive impairment. The results showed that TGA may prevent memory impairment by exerting antioxidant and anti-inflammatory effects in AßO-injected mice. Moreover, TGA exhibited a persistent neuroprotective effect compared to memantine, reflecting an innovative profile of this promising agent against neurodegenerative diseases, such as AD.

The medial prefrontal cortex and the cardiac baroreflex activity: physiological and pathological implications.

The cardiac baroreflex is an autonomic neural mechanism involved in the modulation of the cardiovascular system. It influences the heart rate and peripheral vascular resistance to preserve arterial blood pressure within a narrow variation range. This mechanism is mainly controlled by medullary nuclei located in the brain stem. However, supramedullary areas, such as the ventral portion of medial prefrontal cortex (vMPFC), are also involved. Particularly, the glutamatergic NMDA/NO pathway in the vMPFC can facilitate baroreflex bradycardic and tachycardic responses. In addition, cannabinoid receptors in this same area can reduce or increase those cardiac responses, possibly through alteration in glutamate release. This vMPFC network has been associated to cardiovascular responses during stressful situations. Recent results showed an involvement of glutamatergic, nitrergic, and endocannabinoid systems in the blood pressure and heart rate increases in animals after aversive conditioning. Consequently, baroreflex could be modified by the vMPFC neurotransmission during stressful situations, allowing necessary cardiovascular adjustments. Remarkably, some mental, neurological and neurodegenerative disorders can involve damage in the vMPFC, such as posttraumatic stress disorder, major depressive disorder, Alzheimer's disease, and neuropathic pain. These pathologies are also associated with alterations in glutamate/NO release and endocannabinoid functions along with baroreflex impairment. Thus, the vMPFC seems to play a crucial role on the baroreflex control, either during pathological or physiological stress-related responses. The study of baroreflex mechanism under such pathological view may be helpful to establish causality mechanisms for the autonomic and cardiovascular imbalance found in those conditions. It can explain in the future the reasons of the high cardiovascular risk some neurological and neurodegenerative disease patients undergo. Additionally, the present work offers insights on the possible contributions of vMPFC dysfunction on baroreflex alterations, which, in turn, may raise questions in what extent other brain areas may play a role in autonomic deregulation under such pathological situations.

Distinct sex-dependent behavioral responses induced by two positive allosteric modulators of alpha 5 subunit-containing GABAA receptors.

Dysregulation of GABAergic neurotransmission has long been implicated in several psychiatric disorders, including schizophrenia, depression, and anxiety disorders. Alpha 5 subunit-containing GABAA receptors (α5-GABAAR), which are expressed mainly by pyramidal neurons in the hippocampus, have been proposed as a potential target to treat these psychiatric disorders. Here, we evaluated the effects produced by GL-II-73 and SH-053-2'F-R-CH3 (1, 5, and 10 mg/kg), two positive allosteric modulators of α5-GABAAR in behavioral tests sensitive to drugs with anxiolytic, antidepressant, and antipsychotic properties in male and female C57BL/6 mice. In both males and females, GL-II-73 produced an anxiolytic-like effect in the elevated plus-maze (EPM) and novelty-suppressed feeding and a rapid and sustained antidepressant-like effect in the forced swim test. GL-II-73 also induced antipsychotic-like effects in males indicated by attenuating MK-801-induced hyperlocomotion and prepulse inhibition (PPI) disruption. However, GL-II-73 per se increased locomotor activity and impaired fear memory extinction in males and females and PPI in males. On the other hand, SH-053-2'F-R-CH3 induced anxiolytic-like effects in the EPM and facilitated fear memory extinction in males. Contrary to GL-II-73, SH-053-2'F-R-CH3 attenuated MK-801-induced hyperlocomotion and PPI disruption in females but not in males. Neither of these drugs induced rewarding effects or impaired motor coordination. These findings suggest that GL-II-73 and SH-053-2'F-R-CH3 cause distinct sex-dependent behavioral responses and support continued preclinical research on the potential of positive allosteric modulators of α5-GABAAR for the treatment of psychiatric disorders.

Ventromedial prefrontal cortex CRF1 receptors modulate the tachycardic activity of baroreflex.

Ventral medial prefrontal cortex (vMPFC) glutamatergic neurotransmission has a facilitatory role on cardiac baroreflex activity which is mediated by NMDA receptors activation. Corticotrophin releasing factor receptors type1 and 2 (CRF1 and CRF2), present in the vMPFC, are colocalized in neurons containing glutamate vesicles, suggesting that such receptors may be involved in glutamate release in this cortical area. Therefore, our hypothesis is that the CRF1 and CRF2 receptors can modulate the baroreflex bradycardic and tachycardic responses. In order to prove this assumption, male Wistar rats had bilateral stainless steel guide cannula implanted into the vMPFC, and baroreflex was activated by intravenous infusion of phenylephrine or sodium nitroprusside through a vein catheter. A second catheter was implanted into the femoral artery for cardiovascular measurements. The CRF1 receptor antagonist administration in either infralimbic cortex (IL) or prelimbic cortex (PL), vMPFC regions, was unable to change the bradycardic responses but increased the slope of the baroreflex tachycardic activity. Microinjection of the CRF2 receptor antagonist into the IL and PL did not alter ether bradycardic nor tachycardic baroreflex responses. The administration of the non-selective CRF receptors agonist, urocortin in these areas, did not modify bradycardic responses but decreased tachycardia slope of the baroreflex. CRF1 receptor antagonist administration prior to non-selective CRF agonist in vMPFC prevented the tachycardic responses reduction. However, CRF2 receptor antagonism could not prevent the effect of CRF receptors agonist. These results suggest that IL and PL CRF1 but not CRF2 receptors have an inhibitory role on the baroreflex tachycardic activity. Furthermore, they have no influence on baroreflex bradycardic activity.

κ-Opioid receptors in the medial amygdaloid nucleus modulate autonomic and neuroendocrine responses to acute stress.

The medial amygdaloid nucleus (MeA) is a key neural structure in triggering physiologic and behavioral control during aversive situations. However, MeA role during stress exposure has not yet been fully elucidated. Thus, in the present study, we investigated the involvement of the MeA opioid neurotransmission in the modulation of autonomic, neuroendocrine and behavioral responses evoked by acute restraint stress (RS). The bilateral microinjection of naloxone (non-selective opioid antagonist) into the MeA potentiated RS-evoked autonomic responses and increased plasma corticosterone levels, in a dose-dependent manner. However, no effects were observed in RS-evoked increases on plasma oxytocin levels and anxiogenic-like behavior. Similar to naloxone, MeA pretreatment with the selective κ-opioid antagonist (nor-BNI) also enhanced heart rate and corticosterone increases induced by RS, whereas treatment with selective µ- or δ-opioid antagonists did not affect the physiologic and behavioral responses caused by RS. The present results showed MeA κ-opioid receptors modulate heart rate and corticosterone increases evoked by acute RS, reinforcing the idea of an inhibitory role exerted by MeA during aversive situations .

Bed nucleus of the stria terminalis modulates baroreflex cardiac activity: an interaction between alpha-1 receptors and NMDA/nitric oxide pathway.

The bed nucleus of the stria terminalis (BNST) is a forebrain structure, involved in the modulation of neuroendocrine, cardiovascular and autonomic responses. One of the responses is baroreflex activity, which consists in a neural mechanism responsible for keeping the blood pressure within a narrow range of variation. It has been reported that blockade of BNST α1-adrenoceptors increased the bradycardic component of baroreflex. In addition, such receptors are able to modulate glutamate release in this structure. Interestingly, BNST NMDA receptor antagonism and neuronal nitric oxide synthase (nNOS) inhibition led to the same effect of the α1-adrenoceptors blockade on baroreflex bradycardic response. Therefore, the hypothesis of the present study is that BNST noradrenergic transmission interacts with NMDA/NO pathway through α1 adrenoceptors to modulate the baroreflex activity. Male Wistar rats had stainless steel guide cannulas bilaterally implanted in the BNST. Subsequently, a catheter was inserted into the femoral artery for cardiovascular recordings, and into the femoral vein for assessing baroreflex activation. Injection of the noradrenaline reuptake inhibitor reboxetine in the BNST did not modify the tachycardic, but significantly decreased the bradycardic component of baroreflex. Administration of an α1, but not an α2 antagonist into the BNST prior to reboxetine prevented this effect. Likewise, previous injection of NMDA/NO pathway blockers inhibited the effect of reboxetine on bradycardic response. In conclusion, it was demonstrated for the first time the existence of an interaction between BNST noradrenergic, glutamatergic and nitrergic neurotransmissions in the modulation of bradycardic baroreflex response.

Nitric oxide in the insular cortex modulates baroreflex responses in a cGMP-independent pathway.

Insular cortex is a brain structure involved in the modulation of autonomic activity and cardiovascular function. The nitric oxide/cyclic guanosine-3',5'-monophosphate pathway is a prominent signaling mechanism in the central nervous system, controlling behavioral and physiological responses. Nevertheless, despite evidence regarding the presence of nitric oxide-synthesizing neurons in the insular cortex, its role in the control of autonomic and cardiovascular function has never been reported. Thus, the present study aimed to investigate the involvement of nitric oxide/cyclic guanosine-3',5'-monophosphate pathway mediated by neuronal nitric oxide synthase (nNOS) activation within the insular cortex in the modulation of baroreflex responses in unanesthetized rats. For this, we evaluated the effect of bilateral microinjection of either the nitric oxide scavenger carboxy-PTIO, the selective neuronal nitric oxide synthase inhibitor Nω-Propyl-l-arginine or the soluble guanylate cyclase inhibitor ODQ into the insular cortex on the bradycardia evoked by blood pressure increases in response to intravenous infusion of phenylephrine, and the tachycardia caused by blood pressure decreases evoked by intravenous infusion of sodium nitroprusside. Bilateral microinjection of either NPLA or carboxy-PTIO into the insular cortex increased the reflex bradycardic response, whereas the reflex tachycardia was decreased by these treatments. Bilateral microinjection of the soluble guanylate cyclase inhibitor into the insular cortex did not affect any parameter of baroreflex function evaluated. Overall, our findings provide evidence that insular cortex nitrergic signaling, acting via neuronal nitric oxide synthase, plays a prominent role in control of baroreflex function. However, control of reflex responses seems to be independent of soluble guanylate cyclase activation.

Nitrergic neurotransmission in the paraventricular nucleus of the hypothalamus modulates autonomic, neuroendocrine and behavioral responses to acute restraint stress in rats.

We investigated the involvement of nitrergic neurotransmission within the paraventricular nucleus of the hypothalamus (PVN) in modulation of local neuronal activation, autonomic and neuroendocrine responses and behavioral consequences of acute restraint stress in rats. Bilateral microinjections of the selective neuronal nitric oxide (NO) synthase (nNOS) inhibitor Nw-Propyl-L-arginine (NPLA) or the NO scavenger carboxy-PTIO into the PVN reduced arterial pressure and heart rate increases, as well as the fall in cutaneous tail temperature induced by restraint stress. PVN injection of either NPLA or carboxy-PTIO also inhibited restraint-induced increases in anxiety-related behaviors in the elevated plus-maze 24 h later. Local microinjection of NPLA or carboxy-PTIO into the PVN reduced the number of c-fos-immunoreactive neurons in the dorsal parvocellular, ventromedial, medial parvocellular and lateral magnocelllular portions of the PVN in animals subjected to restraint stress. However, neither NPLA nor carboxy-PTIO into the PVN affected restraint-induced increases in plasma corticosterone concentration. The present results indicate that PVN nitrergic neurotransmission acting via nNOS activation has a facilitatory influence on autonomic responses to acute restraint and the delayed emotional consequences of restraint stress. Our results also provide evidence of a prominent role of local nitrergic neurotransmission in PVN neuronal activation during stress.

Fear extinction disruption in a developmental rodent model of schizophrenia correlates with an impairment in basolateral amygdala-medial prefrontal cortex plasticity.

Schizophrenia patients typically exhibit prominent negative symptoms associated with deficits in extinction recall and decreased ventromedial prefrontal cortex activity (vmPFC, analogous to medial PFC infralimbic segment in rodents). mPFC activity modulates the activity of basolateral amygdala (BLA) and this connectivity is related to extinction. mPFC and BLA activity has been shown to be altered in the methylazoxymethanol acetate (MAM) developmental disruption model of schizophrenia. However, it is unknown if there are alterations in extinction processes in this model. Therefore, we investigated extinction and the role of mPFC-BLA balance in MAM rats. Male offspring of pregnant rats treated with Saline or MAM (20 mg/kg; i.p.) on gestational day 17 were used in fear conditioning (contextual/tone) and electrophysiological experiments (mPFC-BLA plasticity). No difference was observed in conditioning, extinction, and test sessions in contextual fear conditioning. However, MAM-treated rats demonstrated impairment in extinction learning and recall in tone fear conditioning. Furthermore, high frequency stimulation (HFS) of the BLA decreased spike probability in the mPFC of saline-treated rats but not in MAM rats. NMDA antagonist microinjected into the BLA disrupted extinction learning and recall in control rats, resulting in a similar deficit as that observed in MAM-treated rats. These data demonstrate extinction impairment in the MAM model that is analogous to that observed in schizophrenia patients, that was probably due to disruption in the regulation of mPFC activity by glutamatergic neurotransmission in the BLA.

Medial prefrontal cortex TRPV1 and CB1 receptors modulate cardiac baroreflex activity by regulating the NMDA receptor/nitric oxide pathway.

The ventral medial prefrontal cortex (vMPFC) facilitates the cardiac baroreflex response through N-methyl-D-aspartate (NMDA) receptor activation and nitric oxide (NO) formation by neuronal NO synthase (nNOS) and soluble guanylate cyclase (sGC) triggering. Glutamatergic transmission is modulated by the cannabinoid receptor type 1 (CB1) and transient receptor potential vanilloid type 1 (TRPV1) receptors, which may inhibit or stimulate glutamate release in the brain, respectively. Interestingly, vMPFC CB1 receptors decrease cardiac baroreflex responses, while TRPV1 channels facilitate them. Therefore, the hypothesis of the present study is that the vMPFC NMDA/NO pathway is regulated by both CB1 and TRPV1 receptors in the modulation of cardiac baroreflex activity. In order to test this assumption, we used male Wistar rats that had stainless steel guide cannulae bilaterally implanted in the vMPFC. Subsequently, a catheter was inserted into the femoral artery, for cardiovascular recordings, and into the femoral vein for assessing baroreflex activation. The increase in tachycardic and bradycardic responses observed after the microinjection of a CB1 receptors antagonist into the vMPFC was prevented by an NMDA antagonist as well as by the nNOS and sGC inhibition. NO extracellular scavenging also abolished these responses. These same pharmacological manipulations inhibited cardiac reflex enhancement induced by TRPV1 agonist injection into the area. Based on these results, we conclude that vMPFC CB1 and TRPV1 receptors inhibit or facilitate the cardiac baroreflex activity by stimulating or blocking the NMDA activation and NO synthesis.

Neuroreflex control of cardiovascular function is impaired after acute poisoning with chlorpyrifos, an organophosphorus insecticide: Possible short and long term clinical implications.

Although it is well-established that severe poisoning by organophosphorus (OP) compounds strongly affects the cardiorespiratory system, the effects of sub-lethal exposure to these compounds on the neural control of cardiovascular function are poorly explored. The aim of this study was to evaluate the effects of acute sub-lethal exposure to chlorpyrifos (CPF), a commonly used OP insecticide, on three basic reflex mechanisms involved in blood pressure regulation, the peripheral chemoreflex, the baroreflex and the Bezold-Jarisch reflex. Adult male Wistar rats were injected intraperitoneally with a single dose of CPF (30 mg/kg) or saline (0.9%). 24 h after injections, cardiovascular reflexes were tested in awake rats. Potassium cyanide (KCN) and phenylbiguanide (PBG) were injected intravenously to activate the chemoreflex and the Bezold-Jarisch reflex, respectively. The baroreflex was activated by phenylephrine and sodium nitroprusside infusions. Blood samples were taken for measurements of butyrylcholinesterase (BChE) activity while acetylcholinesterase (AChE) activity was measured in brainstem samples. Animals treated with CPF presented signs of intoxication such as ataxia, tremor, lacrimation, salivation, tetany, urination and defecation. The hypertensive and the bradycardic responses of the chemoreflex as well as the hypotensive and bradycardic responses of the Bezold-Jarisch reflex were attenuated in CPF treated animals (P < 0.05). Concerning the baroreflex responses, CPF treatment reduced the bradycardia plateau, the range and the gain of the reflex (P < 0.05). Plasma BChE and brainstem AChE were both reduced significantly after CPF treatment (P < 0.05). Our results showed that acute sub-lethal exposure to CPF impairs the cardiovascular responses of homeostatic and defensive cardiovascular reflexes. These effects are associated with a marked inhibition of plasma BChE and brainstem AChE.

Bed nucleus of the stria terminalis NMDA receptors and nitric oxide modulate contextual fear conditioning in rats.

The bed nucleus of the stria terminalis (BNST) modulates anxiety-like responses, including conditioned emotional responses. Evidence suggests that glutamatergic neurotransmission in the BNST plays a role in the modulation of defensive responses. However, little is known about the involvement of glutamate NMDA receptor activation within the BNST, and its resultant increase in nitric oxide (NO) levels, in the expression of contextual fear conditioning (CFC). We investigated whether the antagonism of NMDA receptors or the reduction of NO levels in the BNST would attenuate behavioral and autonomic responses (i.e. increase in arterial pressure and heart rate, and decrease in tail cutaneous temperature) of rats submitted to a CFC paradigm. Intra-BNST infusion of AP7, an NMDA receptor antagonist, attenuated both behavioral and autonomic changes induced by CFC. Similar results were observed with NPLA and c-PTIO, an nNOS inhibitor and an NO scavenger, respectively. A positive correlation between BNST NO levels and the time spent in freezing behavior was also observed for animals submitted to the CFC. These findings indicate that the expression of CFC involves a facilitation of BNST NMDA receptor-NO signaling. This article is part of the Special Issue entitled 'Ionotropic glutamate receptors'.

Ventrolateral periaqueductal grey matter neurotransmission modulates cardiac baroreflex activity.

Baroreflex activity is a neural mechanism responsible for short-term adjustments in blood pressure (BP). Several supramedullary areas, which send projections to the medulla, are able to control this reflex. In this context, the ventrolateral part of the periaqueductal grey matter (vlPAG), which is a mesencephalic structure, has been suggested to regulate the cardiovascular system. However, its involvement in baroreflex control has never been addressed. Therefore, our hypothesis is that the vlPAG neurotransmission is involved in baroreflex cardiac activity. Male Wistar rats had stainless steel guide cannulae unilaterally or bilaterally implanted in the vlPAG. Afterward, a catheter was inserted into the femoral artery for BP and HR recording. A second catheter was implanted into the femoral vein for baroreflex activation. When the nonselective synaptic blocker cobalt chloride (CoCl2 ) was unilaterally injected into the vlPAG, in either the left or the right hemisphere, it increased the tachycardic response to baroreflex activation. However, when CoCl2 was bilaterally microinjected into the vlPAG it decreased the tachycardic response to baroreflex stimulation. This work shows that vlPAG neurotransmission is involved in modulation of the tachycardic response of the baroreflex. Moreover, we suggest that the interconnections between the vlPAG of both hemispheres are activated during baroreflex stimulation. In this way, our work helps to improve the understanding about brain-heart circuitry control, emphasizing the role of the autonomic nervous system in such modulation.

Prelimbic cortex GABAA receptors are involved in the mediation of restraint stress-evoked cardiovascular responses.

Stress is a response of the organism to homeostasis-threatening stimuli and is coordinated by two main neural systems: the hypothalamic-pituitary-adrenal and the autonomic nervous system. Acute restraint stress (RS) is a model of unavoidable stress, which is characterized by autonomic responses including an increase in mean arterial pressure (MAP) and heart rate (HR), as well as a drop in tail temperature. The prelimbic cortex (PL) has been implicated in the modulation of functional responses caused by RS. The present study aimed to evaluate the role of PL GABAergic neurotransmission in the modulation of autonomic changes induced by RS. Bilateral microinjection of the GABAA receptor antagonist bicuculline methiodide into the PL reduced pressor and tachycardic responses evoked by RS, in a dose-dependent manner, without affecting the tail temperature drop evoked by RS. In order to investigate which peripheral autonomic effector modulated the reduction in RS-cardiovascular responses caused by the blockade of PL GABAA receptors, rats were intravenously pretreated with either atenolol or homatropine methylbromide. The blockade of the cardiac sympathetic nervous system with atenolol blunted the reducing effect of PL treatment with bicuculline methiodide on RS-evoked pressor and tachycardic responses. The blockade of the parasympathetic nervous system with homatropine methylbromide, regardless of affecting the beginning of the tachycardic response, did not impact on the reduction of RS-evoked tachycardic and pressor responses caused by the PL treatment with bicuculline methiodide. The present results indicate that both cardiac sympathetic and parasympathetic activities are involved in the reduction of RS-evoked cardiovascular responses evidenced after the blockade of PL GABAA receptors by bicuculline methiodide.

NOP receptors in the prelimbic cortex have an inhibitory influence on cardiovascular responses induced by restraint stress.

Nociceptin/orphanin FQ (N/OFQ) and its receptor (NOP) have structural homology with classic opioids, but constitute a distinct neurotransmitter system because they lack affinity for the opioid peptides and receptors. This neurotransmission is implicated in several physiologic processes, but the role played by NOP receptors during stress situations remains unclear. The acute restraint stress (RS) is a model of unavoidable stress, characterized by sustained increases in mean arterial pressure (MAP), heart rate (HR) and a drop in tail temperature. On another side, the prelimbic (PL) and infralimbic (IL) cortices, subdivisions of the medial prefrontal cortex (MPFC), are implicated in the modulation of functional responses caused by RS. Considering that, the objective of the present study was to investigate the involvement of PL and IL NOP receptors in the control of autonomic responses induced by RS. Bilateral microinjection of nociceptin (NOP agonist) into the PL reduced the cardiovascular responses evoked by RS. Bilateral microinjection of UPF-101 (NOP antagonist) into the PL potentiated the pressor and tachycardiac responses evoked by RS, in a dose-dependent manner. Local pretreatment with UPF-101 blocked the RS-evoked changes following nociceptin administration into the PL. None of these treatments affected the drop in tail temperature induced by RS. Otherwise, the administration of nociceptin or UPF-101 into the IL had no effect on RS-evoked autonomic changes. To investigate the peripheral mechanism involved in the increase in the RS-evoked cardiovascular responses induced by the blockade of PL NOP receptors, rats were intravenous pretreated with either homatropine or atenolol. The intravenous treatment with homatropine blunted the increase in the RS-evoked pressor and tachycardiac response induced by the PL treatment with UPF-101, while the intravenous treatment with atenolol did not affect the RS-evoked pressor and tachycardiac response induced by the PL treatment with UPF-101. In conclusion, our study shows an influence of the PL N/OFQ neurotransmission, but not the IL NOP receptors, in the control of cardiovascular responses observed during acute stress, by increasing cardiac parasympathetic activity.

Chronic fluoxetine treatment increases NO bioavailability and calcium-sensitive potassium channels activation in rat mesenteric resistance arteries.

Fluoxetine, a selective serotonin reuptake inhibitor (SSRI), has effects beyond its antidepressant properties, altering, e.g., mechanisms involved in blood pressure and vasomotor tone control. Although many studies have addressed the acute impact of fluoxetine on the cardiovascular system, there is a paucity of information on the chronic vascular effects of this SSRI. We tested the hypothesis that chronic fluoxetine treatment enhances the vascular reactivity to vasodilator stimuli by increasing nitric oxide (NO) signaling and activation of potassium (K+) channels. Wistar rats were divided into two groups: (I) vehicle (water for 21 days) or (II) chronic fluoxetine (10 mg/kg/day in the drinking water for 21 days). Fluoxetine treatment increased endothelium-dependent and independent vasorelaxation (analyzed by mesenteric resistance arteries reactivity) as well as constitutive NO synthase (NOS) activity, phosphorylation of eNOS at Serine1177 and NO production, determined by western blot and fluorescence. On the other hand, fluoxetine treatment did not alter vascular expression of neuronal and inducible NOS or guanylyl cyclase (GC). Arteries from fluoxetine-treated rats exhibited increased relaxation to pinacidil. Increased acetylcholine vasorelaxation was abolished by a calcium-activated K+ channel (KCa) blocker, but not by an inhibitor of KATP channels. On the other hand, vascular responses to Bay 41-2272 and 8-bromo-cGMP were similar between the groups. In conclusion, chronic fluoxetine treatment increases endothelium-dependent and independent relaxation of mesenteric resistance arteries by mechanisms that involve increased eNOS activity, NO generation, and KCa channels activation. These effects may contribute to the cardiovascular effects associated with chronic fluoxetine treatment.

Prelimbic cortex 5-HT1A and 5-HT2C receptors are involved in the hypophagic effects caused by fluoxetine in fasted rats.

The regulation of food intake involves a complex interplay between the central nervous system and the activity of organs involved in energy homeostasis. Besides the hypothalamus, recognized as the center of this regulation, other structures are involved, especially limbic regions such as the ventral medial prefrontal cortex (vMPFC). Monoamines, such as serotonin (5-HT), play an important role in appetite regulation. However, the effect in the vMPFC of the selective serotonin reuptake inhibitor (SSRI), fluoxetine, on food intake has not been studied. The aim of the present study was to study the effects on food intake of fed and fasted rats evoked by fluoxetine injection into the prelimbic cortex (PL), a sub-region of the vMPFC, or given systemically, and which 5-HT receptors in the PL are involved in fluoxetine responses. Fluoxetine was injected into the PL or given systemically in male Wistar rats. Independent groups of rats were pretreated with intra-PL antagonists of 5-HT receptors: 5-HT1A (WAY100635), 5-HT2C (SB242084) or 5-HT1B (SB216641). Fluoxetine (0.1; 1; 3; 10nmol/200nL) injected into the PL induced a dose-dependent hypophagic effect in fasted rats. This effect was reversed by prior local treatment with WAY100635 (1; 10nmol) or SB242084 (1; 10nmol), but not with SB216641 (0.2; 2.5; 10nmol). Systemic fluoxetine induced a hypophagic effect, which was blocked by intra-PL 5-HT2C antagonist (10nmol) administration. Our findings suggest that PL 5-HT neurotransmission modulates the central control of food intake and 5-HT1A and 5-HT2C receptors in the PL could be potential targets for the action of fluoxetine.

κ-opioid receptors in the infralimbic cortex modulate the cardiovascular responses to acute stress.

NEW FINDINGS: What is the central question of this study? A brief experience of stress can cause structural remodelling in the infralimbic cortex. In the present study, we addressed the potential role played by opioidergic neurotransmission in the infralimbic cortex in the modulation of stress-evoked autonomic responses. What is the main finding and its importance? Using the restraint stress model, we showed that infralimbic cortex κ-opioid receptors, but not µ- and δ-opioid receptors, modulate stress-evoked cardiovascular responses. The infralimbic cortex (IL) is known to modulate behavioural and physiological responses during aversive situations. We investigated the hypothesis that opioid neurotransmission in the IL modulates the autonomic responses induced in rats subjected to restraint stress (RS). Male Wistar rats (250-280 g) were used. Guide cannulae were implanted bilaterally in the IL for the microinjection of either drugs or vehicle, and a polyethylene catheter was implanted into the femoral artery for recording of mean arterial pressure (MAP) and heart rate (HR) using a computerized acquisition system. Tail temperature was evaluated using a thermal camera. Rats were subjected to RS 10 min after the microinjection of drugs or vehicle into the IL. Exposure to RS evoked hypertension, tachycardia and a reduction in tail temperature. Bilateral microinjections of the non-selective opioid antagonist naloxone into the IL generated an inverted U-shaped dose-inhibition curve on RS-evoked MAP and HR responses. Microinjection of nor-BNI (κ-selective antagonist) reduced the increases in MAP and HR evoked by RS. In contrast, pretreatment of the IL with CTAP (µ-selective antagonist) or naltrindole (δ-selective antagonist) had no effect on the restraint-evoked increases in MAP and HR. None of these treatments altered the reduction in the tail temperature evoked by RS. In conclusion, κ-opioid receptors in the IL modulate pressor and tachycardiac responses caused by RS, suggesting a facilitatory role of this structure in this aversive situation.

Toll-like receptor 9 plays a key role in the autonomic cardiac and baroreflex control of arterial pressure.

The crosstalk between the immune and the autonomic nervous system may impact the cardiovascular function. Toll-like receptors are components of the innate immune system and play developmental and physiological roles. Toll-like receptor 9 (TLR9) is involved in the pathogenesis of cardiovascular diseases, such as hypertension and heart failure. Since such diseases are commonly accompanied by autonomic imbalance and lower baroreflex sensitivity, we hypothesized that TLR9 modulates cardiac autonomic and baroreflex control of arterial pressure (AP). Toll-like receptor 9 knockout (TLR9 KO) and wild-type (WT) mice were implanted with catheters into carotid artery and jugular vein and allowed to recover for 3 days. After basal recording of AP, mice received methyl-atropine or propranolol. AP and pulse interval (PI) variability were evaluated in the time and frequency domain (spectral analysis), as well as by multiscale entropy. Spontaneous baroreflex was studied by sequence technique. Behavioral and cardiovascular responses to fear-conditioning stress were also evaluated. AP was similar between groups, but TLR9 KO mice exhibited lower basal heart rate (HR). AP variability was not different, but PI variability was increased in TLR9 KO mice. The total entropy was higher in TLR9 KO mice. Moreover, baroreflex function was found higher in TLR9 KO mice. Atropine-induced tachycardia was increased in TLR9 KO mice, whereas the propranolol-induced bradycardia was similar to WT mice. TLR9 KO mice exhibit increased behavioral and decreased tachycardia responses to fear-conditioning stress. In conclusion, our findings suggest that TLR9 may negatively modulate cardiac vagal tone and baroreflex in mice.

Increased Contextual Fear Conditioning in iNOS Knockout Mice: Additional Evidence for the Involvement of Nitric Oxide in Stress-Related Disorders and Contribution of the Endocannabinoid System.

BACKGROUND: Inducible or neuronal nitric oxide synthase gene deletion increases or decreases anxiety-like behavior in mice, respectively. Since nitric oxide and endocannabinoids interact to modulate defensive behavior, the former effect could involve a compensatory increase in basal brain nitric oxide synthase activity and/or changes in the endocannabinoid system. Thus, we investigated the expression and extinction of contextual fear conditioning of inducible nitric oxide knockout mice and possible involvement of endocannabinoids in these responses. METHODS: We evaluated the effects of a preferential neuronal nitric oxide synthase inhibitor, 7-nitroindazol, nitric oxide synthase activity, and mRNA changes of nitrergic and endocannabinoid systems components in the medial prefrontal cortex and hippocampus of wild-type and knockout mice. The effects of URB597, an inhibitor of the fatty acid amide hydrolase enzyme, which metabolizes the endocannabinoid anandamide, WIN55,212-2, a nonselective cannabinoid agonist, and AM281, a selective CB1 antagonist, on contextual fear conditioning were also evaluated. RESULTS: Contextual fear conditioning expression was similar in wild-type and knockout mice, but the latter presented extinction deficits and increased basal nitric oxide synthase activity in the medial prefrontal cortex. 7-Nitroindazol decreased fear expression and facilitated extinction in wild-type and knockout mice. URB597 decreased fear expression in wild-type and facilitated extinction in knockout mice, whereas WIN55,212-2 and AM281 increased it in wild-type mice. Nonconditioned knockout mice showed changes in the mRNA expression of nitrergic and endocannabinoid system components in the medial prefrontal cortex and hippocampus that were modified by fear conditioning. CONCLUSION: These data reinforce the involvement of the nitric oxide and endocannabinoids (anandamide) in stress-related disorders and point to a deregulation of the endocannabinoid system in situations where nitric oxide signaling is increased.