Activation of mGluR1 negatively modulates glutamate-induced phase shifts of the circadian pacemaker in the mouse suprachiasmatic nucleus.

In mammals, photic information delivered to the suprachiasmatic nucleus (SCN) via the retinohypothalamic tract (RHT) plays a crucial role in synchronizing the master circadian clock located in the SCN to the solar cycle. It is well known that glutamate released from the RHT terminals initiates the synchronizing process by activating ionotropic glutamate receptors (iGluRs) on retinorecipient SCN neurons. The potential role of metabotropic glutamate receptors (mGluRs) in modulating this signaling pathway has received less attention. In this study, using extracellular single-unit recordings in mouse SCN slices, we investigated the possible roles of the Gq/11 protein-coupled mGluRs, mGluR1 and mGluR5, in photic resetting. We found that mGluR1 activation in the early night produced phase advances in neural activity rhythms in the SCN, while activation in the late night produced phase delays. In contrast, mGluR5 activation had no significant effect on the phase of these rhythms. Interestingly, mGluR1 activation antagonized phase shifts induced by glutamate through a mechanism that was dependent upon CaV1.3 L-type voltage-gated Ca2+ channels (VGCCs). While both mGluR1-evoked phase delays and advances were inhibited by knockout (KO) of CaV1.3 L-type VGCCs, different signaling pathways appeared to be involved in mediating these effects, with mGluR1 working via protein kinase G in the early night and via protein kinase A signaling in the late night. We conclude that, in the mouse SCN, mGluR1s function to negatively modulate glutamate-evoked phase shifts.

Oestrogen inhibits salt-dependent hypertension by suppressing GABAergic excitation in magnocellular AVP neurons.

AIMS: Abundant evidence indicates that oestrogen (E2) plays a protective role against hypertension. Yet, the mechanism underlying the antihypertensive effect of E2 is poorly understood. In this study, we sought to determine the mechanism through which E2 inhibits salt-dependent hypertension. METHODS AND RESULTS: To this end, we performed a series of in vivo and in vitro experiments employing a rat model of hypertension that is produced by deoxycorticosterone acetate (DOCA)-salt treatment after uninephrectomy. We found that E2 prevented DOCA-salt treatment from inducing hypertension, raising plasma arginine-vasopressin (AVP) level, enhancing the depressor effect of the V1a receptor antagonist (Phenylac1,D-Tyr(Et)2,Lys6,Arg8,des-Gly9)-vasopressin, and converting GABAergic inhibition to excitation in hypothalamic magnocellular AVP neurons. Moreover, we obtained results indicating that the E2 modulation of the activity and/or expression of NKCC1 (Cl- importer) and KCC2 (Cl- extruder) underpins the effect of E2 on the transition of GABAergic transmission in AVP neurons. Lastly, we discovered that, in DOCA-salt-treated hypertensive ovariectomized rats, CLP290 (prodrug of the KCC2 activator CLP257, intraperitoneal injections) lowered blood pressure, and plasma AVP level and hyperpolarized GABA equilibrium potential to prevent GABAergic excitation from emerging in the AVP neurons of these animals. CONCLUSION: Based on these results, we conclude that E2 inhibits salt-dependent hypertension by suppressing GABAergic excitation to decrease the hormonal output of AVP neurons.

Excessive maternal salt intake gives rise to vasopressin-dependent salt sensitivity of blood pressure in male offspring.

Salt sensitivity of blood pressure (SSBP) is a trait carrying strong prognostic implications for various cardiovascular diseases. To test the hypothesis that excessive maternal salt intake causes SSBP in offspring through a mechanism dependent upon arginine-vasopressin (AVP), we performed a series of experiments using offspring of the rat dams salt-loaded during pregnancy and lactation with 1.5% saline drink ("experimental offspring") and those with normal perinatal salt exposure ("control offspring"). Salt challenge, given at 7-8 weeks of age with either 2% saline drink (3 days) or 8% NaCl-containing chow (4 weeks), had little or no effect on systolic blood pressure (SBP) in female offspring, whereas the salt challenge significantly raised SBP in male offspring, with the magnitude of increase being greater in experimental, than control, rats. Furthermore, the salt challenge not only raised plasma AVP level more and caused greater depressor responses to V1a and V2 AVP receptor antagonists to occur in experimental, than control, males, but it also made GABA excitatory in a significant proportion of magnocellular AVP neurons of experimental males by depolarizing GABA equilibrium potential. The effect of the maternal salt loading on the salt challenge-elicited SBP response in male offspring was precluded by maternal conivaptan treatment (non-selective AVP receptor antagonist) during the salt-loading period, whereas it was mimicked by neonatal AVP treatment. These results suggest that the excessive maternal salt intake brings about SSBP in male offspring, both the programming and the expression of which depend on increased AVP secretion that may partly result from excitatory GABAergic action.

Role of T-type Calcium Channels in Generating Hyperexcitatory Behaviors during Emergence from Sevoflurane Anesthesia in Neonatal Rats.

In the current study, we sought to investigate whether T-type Ca2+ channels (TCCs) in the brain are involved in generating post-anesthetic hyperexcitatory behaviors (PAHBs). We found that younger rat pups (postnatal days 9-11) had a higher incidence of PAHBs and higher PAHB scores than older pups (postnatal days 16-18) during emergence from sevoflurane anesthesia. The power spectrum of the theta oscillations (4 Hz-8 Hz) in the prefrontal cortex was significantly enhanced in younger pups when PAHBs occurred, while there were no significant changes in older pups. Both the power of theta oscillations and the level of PAHBs were significantly reduced by the administration of TCC inhibitors. Moreover, the sensitivity of TCCs in the medial dorsal thalamic nucleus to sevoflurane was found to increase with age by investigating the kinetic properties of TCCs in vitro. TCCs were activated by potentiated GABAergic depolarization with a sub-anesthetic dose of sevoflurane (1%). These data suggest that (1) TCCs in the brain contribute to the generation of PAHBs and the concomitant electroencephalographic changes; (2) the stronger inhibitory effect of sevoflurane contributes to the lack of PAHBs in older rats; and (3) the contribution of TCCs to PAHBs is not mediated by a direct effect of sevoflurane on TCCs.

Long-term ionic plasticity of GABAergic signalling in the hypothalamus.

The hypothalamus contains a number of nuclei that subserve a variety of functions, including generation of circadian rhythms, regulation of hormone secretion and maintenance of homeostatic levels for a variety of physiological parameters. Within the hypothalamus, γ-amino-butyric acid (GABA) is one of the major neurotransmitters responsible for cellular communication. Although GABA most commonly serves as an inhibitory neurotransmitter, a growing body of evidence indicates that it can evoke post-synaptic excitation as a result of the active regulation of intracellular chloride concentration. In this review, we consider the evidence for this ionic plasticity of GABAergic synaptic transmission in five distinct cases in hypothalamic cell populations. We argue that this plasticity serves as part of the functional response to or is at least associated with dehydration, lactation, hypertension and stress. As such, GABA excitation should be considered as part of the core homeostatic mechanisms of the hypothalamus.

Activation of peripheral group III metabotropic glutamate receptors inhibits pain transmission by decreasing neuronal excitability in the CFA-inflamed knee joint.

Peripheral group III metabotropic glutamate receptors (mGluRs) function to modulate pain signaling in inflammatory states. Here, we established in vivo experimental settings, including dynamic weight bearing test and in vivo single nerve recording, to elucidate how the group III mGluRs contribute to inhibiting pain transmission at the peripheral sensory nerve terminal in inflammatory states (1 and 3 days) elicited by Complete Freund's Adjuvant (CFA). As a result, CFA-induced nociceptive behaviors were significantly alleviated after administration of 100 and 200 µM L-AP4 (l-2-amino-4-phosphonobutylate; group III mGluR agonist). In addition, neuronal discharges evoked by 6- and 26-g von Frey filaments at the nerve significantly decreased after administration of 200 µM L-AP4. However, this event was not observed in non-inflammatory state. These results suggest that the group III mGluRs negatively regulate nociceptive behavior and pain transmission by lessening neuronal firing rates at the peripheral nerve in inflammation.

Long-Term Isolation Elicits Depression and Anxiety-Related Behaviors by Reducing Oxytocin-Induced GABAergic Transmission in Central Amygdala.

Isolation stress is a major risk factor for neuropsychiatric disorders such as depressive and anxiety disorders. However, the molecular mechanisms underlying isolation-induced neuropsychiatric disorders remain elusive. In the present study, we investigated the subcellular mechanisms by which long-term isolation elicits depression and anxiety-related behaviors in mice. First, we found that long-term isolation induced depression-related behaviors in the forced swimming test (FST) and the sucrose preference test, as well as anxiety-related behaviors in the elevated zero maze test (EZMT) and the open field test. Next, we showed that intracentral amygdala (CeA) injection of oxytocin (OXT), but not intracerebroventricular injection, attenuated isolation-induced depression and anxiety-related behaviors via oxytocin receptor (OXTR), not vasopressin-1a receptor (V1aR), in the FST and EZMT, respectively. Quantitative real-time polymerase chain reaction analysis revealed that after 5 weeks of isolation, mRNA transcription of OXTR in the CeA, but not that of V1aR, significantly decreased, whereas OXT and vasopressin mRNA transcription in the paraventricular nucleus of hypothalamus did not change significantly. Whole-cell patch clamping of acute brain slices demonstrated that the frequency of miniature inhibitory postsynaptic currents (mIPSCs) in CeA neurons, but not their amplitude, was lower in isolated mice than in group-housed mice. Notably, OXT treatment increased the mIPSC frequency in the CeA neurons, but to a lesser extent in the case of isolated mice than in that of group-housed mice via OXTR. Taken together, our findings suggest that long-term isolation down-regulates OXTR mRNA transcription and diminishes OXT-induced inhibitory synaptic transmission in the CeA and may contribute to the development of depression and anxiety-related behaviors in isolated mice through the enhancement of CeA activity.

Defined Conditions for Differentiation of Functional Retinal Ganglion Cells From Human Pluripotent Stem Cells.

Purpose: We aimed to establish an efficient method for retinal ganglion cell (RGC) differentiation from human pluripotent stem cells (hPSCs) using defined factors. Methods: To define the contribution of specific signal pathways to RGC development and optimize the differentiation of hPSCs toward RGCs, we examined RGC differentiation in three stages: (1) eye field progenitors expressing the eye field transcription factors (EFTFs), (2) RGC progenitors expressing MATH5, and (3) RGCs expressing BRN3B and ISLET1. By monitoring the condition that elicited the highest yield of cells expressing stage-specific markers, we determined the optimal concentrations and combinations of signaling pathways required for efficient generation of RGCs from hPSCs. Results: Precise modulation of signaling pathways, including Wnt, insulin growth factor-1, and fibroblast growth factor, in combination with mechanical isolation of neural rosette cell clusters significantly enriched RX and PAX6 double-positive eye field progenitors from hPSCs by day 12. Furthermore, Notch signal inhibition facilitated differentiation into MATH5-positive progenitors at 90% efficiency by day 20, and these cells further differentiated to BRN3B and ISLET1 double-positive RGCs at 45% efficiency by day 40. RGCs differentiated via this method were functional as exemplified by their ability to generate action potentials, express microfilament components on neuronal processes, and exhibit axonal transportation of mitochondria. Conclusions: This protocol offers highly defined culture conditions for RGC differentiation from hPSCs and in vitro disease model and cell source for transplantation for diseases related to RGCs.

Differential effects on sodium current impairments by distinct SCN1A mutations in GABAergic neurons derived from Dravet syndrome patients.

BACKGROUND: We investigated how two distinct mutations in SCN1A differentially affect electrophysiological properties of the patient-derived GABAergic neurons and clinical severities in two Dravet syndrome (DS) patients. MATERIALS AND METHODS: We established induced pluripotent stem cells from two DS patients with different mutations in SCN1A and subsequently differentiated them into forebrain GABAergic neurons. Functionality of differentiated GABAergic neurons was examined by electrophysiological recordings. RESULTS: DS-1 patient had a missense mutation, c.4261G > T [GenBank: NM_006920.4] and DS-2 patient had a nonsense frameshift mutation, c.3576_3580 del TCAAA [GenBank: NM_006920.4]. Clinically, contrary to our expectations, DS-1 patient had more severe symptoms including frequency of seizure episodes and the extent of intellectual ability penetration than DS-2 patient. Electrophysiologic recordings showed significantly lower sodium current density and reduced action potential frequency at strong current injection (>60 pA) in GABAergic neurons derived from both. Intriguingly, unique genetic alterations of SCN1A differentially impacted electrophysiological impairment of the neurons, and the impairment's extent corresponded with the symptomatic severity of the donor from which the iPSCs were derived. CONCLUSION: Our results suggest the possibility that patient-derived iPSCs may provide a reliable in vitro system that reflects clinical severities in individuals with DS.

Excitatory GABAergic Action and Increased Vasopressin Synthesis in Hypothalamic Magnocellular Neurosecretory Cells Underlie the High Plasma Level of Vasopressin in Diabetic Rats.

Diabetes mellitus (DM) is associated with increased plasma levels of arginine-vasopressin (AVP), which may aggravate hyperglycemia and nephropathy. However, the mechanisms by which DM may cause the increased AVP levels are not known. Electrophysiological recordings in supraoptic nucleus (SON) slices from streptozotocin (STZ)-induced DM rats and vehicle-treated control rats revealed that γ-aminobutyric acid (GABA) functions generally as an excitatory neurotransmitter in the AVP neurons of STZ rats, whereas it usually evokes inhibitory responses in the cells of control animals. Furthermore, Western blotting analyses of Cl- transporters in the SON tissues indicated that Na+-K+-2Cl- cotransporter isotype 1 (a Cl- importer) was upregulated and K+-Cl- cotransporter isotype 2 (KCC2; a Cl- extruder) was downregulated in STZ rats. Treatment with CLP290 (a KCC2 activator) significantly lowered blood AVP and glucose levels in STZ rats. Last, investigation that used rats expressing an AVP-enhanced green fluorescent protein fusion gene showed that AVP synthesis in AVP neurons was much more intense in STZ rats than in control rats. We conclude that altered Cl- homeostasis that makes GABA excitatory and enhanced AVP synthesis are important changes in AVP neurons that would increase AVP secretion in DM. Our data suggest that Cl- transporters in AVP neurons are potential targets of antidiabetes treatments.

The contribution of activated peripheral kappa opioid receptors (kORs) in the inflamed knee joint to anti-nociception.

The systemic administration of opioids can be used for their strong analgesic effect. However, extensive activation of opioid receptors (ORs) beyond the targeted tissue can cause dysphoria, pruritus, and constipation. Therefore, selective activation of peripheral ORs present in the afferent fibers of the targeted tissue can be considered a superior strategy in opioid analgesia to avoid potential adverse effects. The purpose of this study was to clarify the role of peripheral kappa opioid receptors (kORs) in arthritic pain for the possible use of peripheral ORs as a target in anti-nociceptive therapy. We administered U50488 or nor-BNI/DIPPA, a selective agonist or antagonist of kOR, respectively into arthritic rat knee joints induced using 1% carrageenan. After the injection of U50488 or U50488 with nor-BNI or DIPPA into the inflamed knee joint, we evaluated nociceptive behavior as indicated by reduced weight-bearing on the ipsilateral limbs of the rat and recorded the activity of mechanosensitive afferents (MSA). In the inflamed knee joint, the intra-articular application of 1µM, 10nM, or 0.1nM U50488 resulted in a significant reduction in nociceptive behavior. In addition, 1µM and 10nM U50488 decreased MSA activity. However, in a non-inflamed knee joint, 1µM U50488 had no effect on MSA activity. Additionally, intra-articular pretreatment with 20µM nor-BNI or 10µM DIPPA significantly blocked the inhibitory effects of 1µM U50488 on nociceptive behavior and MSA activity in the inflamed knee joint. These results implicate that peripheral kORs can contribute to anti-nociceptive processing in an inflamed knee joint.

A combination of small molecules directly reprograms mouse fibroblasts into neural stem cells.

The generation of induced neural stem cells (iNSCs) from somatic cells using defined factors provides new avenues for basic research and cell therapies for various neurological diseases, such as Parkinson's disease, Huntington's disease, and spinal cord injuries. However, the transcription factors used for direct reprogramming have the potential to cause unexpected genetic modifications, which limits their potential application in cell therapies. Here, we show that a combination of four chemical compounds resulted in cells directly acquiring a NSC identity; we termed these cells chemically-induced NSCs (ciNSCs). ciNSCs expressed NSC markers (Pax6, PLZF, Nestin, Sox2, and Sox1) and resembled NSCs in terms of their morphology, self-renewal, gene expression profile, and electrophysiological function when differentiated into the neuronal lineage. Moreover, ciNSCs could differentiate into several types of mature neurons (dopaminergic, GABAergic, and cholinergic) as well as astrocytes and oligodendrocytes in vitro. Taken together, our results suggest that stably expandable and functional ciNSCs can be directly reprogrammed from mouse fibroblasts using a combination of small molecules without any genetic manipulation, and will provide a new source of cells for cellular replacement therapy of neurodegenerative diseases.

Histamine 1 receptor-Gßγ-cAMP/PKA-CFTR pathway mediates the histamine-induced resetting of the suprachiasmatic circadian clock.

BACKGROUND: Recent evidence indicates that histamine, acting on histamine 1 receptor (H1R), resets the circadian clock in the mouse suprachiasmatic nucleus (SCN) by increasing intracellular Ca(2+) concentration ([Ca(2+)]i) through the activation of CaV1.3 L-type Ca(2+) channels and Ca(2+)-induced Ca(2+) release from ryanodine receptor-mediated internal stores. RESULTS: In the current study, we explored the underlying mechanisms with various techniques including Ca(2+)- and Cl(-)-imaging and extracellular single-unit recording. Our hypothesis was that histamine causes Cl(-) efflux through cystic fibrosis transmembrane conductance regulator (CFTR) to elicit membrane depolarization needed for the activation of CaV1.3 Ca(2+) channels in SCN neurons. We found that histamine elicited Cl(-) efflux and increased [Ca(2+)]i in dissociated mouse SCN cells. Both of these events were suppressed by bumetanide [Na(+)-K(+)-2Cl(-) cotransporter isotype 1 (NKCC1) blocker], CFTRinh-172 (CFTR inhibitor), gallein (Gßγ protein inhibitor) and H89 [protein kinase A (PKA) inhibitor]. By itself, H1R activation with 2-pyridylethylamine increased the level of cAMP in the SCN and this regulation was prevented by gallein. Finally, histamine-evoked phase shifts of the circadian neural activity rhythm in the mouse SCN slice were blocked by bumetanide, CFTRinh-172, gallein or H89 and were not observed in NKCC1 or CFTR KO mice. CONCLUSIONS: Taken together, these results indicate that histamine recruits the H1R-Gßγ-cAMP/PKA pathway in the SCN neurons to activate CaV1.3 channels through CFTR-mediated Cl(-) efflux and ultimately to phase-shift the circadian clock. This pathway and NKCC1 may well be potential targets for agents designed to treat problems resulting from the disturbance of the circadian system.

The effects of Chamaecyparis obtusa essential oil on pain-related behavior and expression of pro-inflammatory cytokines in carrageenan-induced arthritis in rats.

Chamaecyparis obtusa essential oil (COE) has been widely used to treat allergic diseases and was suggested to exert anti-inflammatory, antioxidant, and antimicrobial effects. This study evaluated the effects of COE on pain-related behavior and pro-inflammatory cytokines in rats with carrageenan (CGN)-induced arthritis. Reduced dynamic weight load on inflamed joint in voluntarily walking rats was used as the behavior test for arthritic pain; 10% COE-treated group was significantly attenuated pain (6-8 h post-CGN injection) compared to VEH (mineral oil)-treated group. In addition, the protein levels of interleukin (IL)-1ß, tumor necrosis factor-α, IL-6 (6-8 h), and cyclooxygenase (COX)-2 (8 h) within the synovial membrane, as well as IL-1ß, COX-2 (6-8 h), and IL-6 (5-7 h) within the meniscus, of 10% COE-treated group were significantly reduced. The current results implicate that COE has anti-inflammatory and anti-nociceptive effects on arthritis in rats.

Histamine resets the circadian clock in the suprachiasmatic nucleus through the H1R-CaV 1.3-RyR pathway in the mouse.

Histamine, a neurotransmitter/neuromodulator implicated in the control of arousal state, exerts a potent phase-shifting effect on the circadian clock in the rodent suprachiasmatic nucleus (SCN). In this study, the mechanisms by which histamine resets the circadian clock in the mouse SCN were investigated. As a first step, Ca(2+) -imaging techniques were used to demonstrate that histamine increases intracellular Ca(2+) concentration ([Ca(2+) ]i ) in acutely dissociated SCN neurons and that this increase is blocked by the H1 histamine receptor (H1R) antagonist pyrilamine, the removal of extracellular Ca(2+) and the L-type Ca(2+) channel blocker nimodipine. The histamine-induced Ca(2+) transient is reduced, but not blocked, by application of the ryanodine receptor (RyR) blocker dantrolene. Immunohistochemical techniques indicated that CaV 1.3 L-type Ca(2+) channels are expressed mainly in the somata of SCN cells along with the H1R, whereas CaV 1.2 channels are located primarily in the processes. Finally, extracellular single-unit recordings demonstrated that the histamine-elicited phase delay of the circadian neural activity rhythm recorded from SCN slices is blocked by pyrilamine, nimodipine and the knockout of CaV 1.3 channel. Again, application of dantrolene reduced but did not block the histamine-induced phase delays. Collectively, these results indicate that, to reset the circadian clock, histamine increases [Ca(2+) ]i in SCN neurons by activating CaV 1.3 channels through H1R, and secondarily by causing Ca(2+) -induced Ca(2+) release from RyR-mediated internal stores.

Conversion of mouse fibroblasts into cardiomyocyte-like cells using small molecule treatments.

The possibility of controlling cell fates by overexpressing specific transcription factors has led to numerous studies in stem cell research. Small molecules can be used, instead of transcription factors, to induce the de-differentiation of somatic cells or to induce pluripotent cells (iPSCs). Here we reported that combinations of small molecules could convert mouse fibroblasts into cardiomyocyte-like cell without requiring transcription factor expression. Treatment with specific combinations of small molecules that are enhancer for iPSC induction converted mouse fibroblasts into spontaneously contracting, cardiac troponin T-positive, cardiomyocyte-like cells. We specifically identified five small molecules that can induce mouse fibroblasts to form these cardiomyocyte-like cells. These cells are similar to primary cardiomyocytes in terms of marker gene expression, epigenetic status of cardiac-specific genes, and subcellular structure. Our findings indicate that lineage conversion can be induced not only by transcription factors, but also by small molecules.

Possible role of GABAergic depolarization in neocortical neurons in generating hyperexcitatory behaviors during emergence from sevoflurane anesthesia in the rat.

Hyperexcitatory behaviors occurring after sevoflurane anesthesia are of serious clinical concern, but the underlying mechanism is unknown. These behaviors may result from the potentiation by sevoflurane of GABAergic depolarization/excitation in neocortical neurons, cells implicated in the genesis of consciousness and arousal. The current study sought to provide evidence for this hypothesis with rats, the neocortical neurons of which are known to respond to GABA (γ-aminobutyric acid) with depolarization/excitation at early stages of development (i.e., until the second postnatal week) and with hyperpolarization/inhibition during adulthood. Employing behavioral tests and electrophysiological recordings in neocortical slice preparations, we found: (1) sevoflurane produced PAHBs (post-anesthetic hyperexcitatory behaviors) in postnatal day (P)1-15 rats, whereas it failed to elicit PAHBs in P16 or older rats; (2) GABAergic PSPs (postsynaptic potentials) were depolarizing/excitatory in the neocortical neurons of P5 and P10 rats, whereas mostly hyperpolarizing/inhibitory in the cells of adult rats; (3) at P14-15, <50% of rats had PAHBs and, in general, the cells of the animals with PAHBs exhibited strongly depolarizing GABAergic PSPs, whereas those without PAHBs showed hyperpolarizing or weakly depolarizing GABAergic PSPs; (4) bumetanide [inhibitor of the Cl- importer NKCC (Na+-K+-2Cl- cotransporter)] treatment at P5 suppressed PAHBs and depolarizing GABAergic responses; and (5) sevoflurane at 1% (i.e., concentration<1 minimum alveolar concentration) potentiated depolarizing GABAergic PSPs in the neurons of P5 and P10 rats and of P14-15 animals with PAHBs, evoking action potentials in ≥50% of these cells. On the basis of these results, we conclude that sevoflurane may produce PAHBs by potentiating GABAergic depolarization/excitation in neocortical neurons.

GABAergic excitation of vasopressin neurons: possible mechanism underlying sodium-dependent hypertension.

RATIONALE: Increased arginine-vasopressin (AVP) secretion is a key physiological response to hyperosmotic stress and may be part of the mechanism by which high-salt diets induce or exacerbate hypertension. OBJECTIVE: Using deoxycorticosterone acetate-salt hypertension model rats, we sought to test the hypothesis that changes in GABA(A) receptor-mediated inhibition in AVP-secreting magnocellular neurons contribute to the generation of Na(+)-dependent hypertension. METHODS AND RESULTS: In vitro gramicidin-perforated recordings in the paraventricular and supraoptic nuclei revealed that the GABAergic inhibition in AVP-secreting neurons was converted into excitation in this model, because of the depolarization of GABA equilibrium potential. Meanwhile, in vivo extracellular recordings in the supraoptic nuclei showed that the GABAergic baroreflexive inhibition of magnocellular neurons was transformed to excitation, so that baroreceptor activation may increase AVP release. The depolarizing GABA equilibrium potential shift in AVP-secreting neurons occurred progressively over weeks of deoxycorticosterone acetate-salt treatment along with gradual increases in plasma AVP and blood pressure. Furthermore, the shift was associated with changes in chloride transporter expression and partially reversed by bumetanide (Na(+)-K(+)-2Cl(-) cotransporter inhibitor). Intracerebroventricular bumetanide administration during deoxycorticosterone acetate-salt treatment hindered the development of hypertension and rise in plasma AVP level. Muscimol (GABA(A) agonist) microinjection into the supraoptic nuclei in hypertensive rats increased blood pressure, which was prevented by previous intravenous V1a AVP antagonist injection. CONCLUSIONS: We conclude that the inhibitory-to-excitatory switch of GABAA receptor-mediated transmission in AVP neurons contributes to the generation of Na(+)-dependent hypertension by increasing AVP release. We speculate that normalizing the GABA equilibrium potential may have some utility in treating Na(+)-dependent hypertension.

Peripheral group II and III metabotropic glutamate receptors in the knee joint attenuate carrageenan-induced nociceptive behavior in rats.

This study sought to evaluate whether peripheral group II and III metabotropic glutamate receptors (mGluRs) in the knee joint have inhibitory effects on carrageenan-induced nociceptive behavior. To this end, changes in weight load and hind paw withdrawal threshold were measured in rats following the administration of specific peripheral group II and III mGluR agonists 30min before (induction phase) and 4h after (maintenance phase) the injection of carrageenan (1%, 50µl). During the induction phase of arthritic pain, a significant recovery of reduced weight load occurred after the administration of 500µM APDC ((2R, 4R)-4-aminopyrrolidine-2,4-dicarboxylate; group II agonist) and 100 and 500µM L-AP4 (l-2-amino-4-phosphonobutylate; group III agonist). Similarly, 100 and 500µM APDC and 500µM L-AP4 significantly reduced mechanical hyperalgesia during the induction phase. In the maintenance phase, APDC at all doses (10, 100 and 500µM) and 100 and 500µM L-AP4 significantly counteracted the reduction in weight load, and APDC and L-AP4 at all doses (10, 100 and 500µM) inhibited mechanical hyperalgesia. The current study suggests that peripheral group II and III mGluRs in the knee joint negatively modulates nociceptive behavior during both the induction and maintenance phases of carrageenan-induced arthritic pain.