GABA consumption during early pregnancy impairs endometrial receptivity and embryo development in mice.

γ-Aminobutyrate (GABA) is commonly used as a food supplement and a health care product by young females, due to its positive roles in relieving stress, alleviating anxiety, and improving sleep. However, its recommended daily dose in different products varies widely. Besides, it is unknown whether, and how, GABA consumption during early pregnancy influences pregnancy establishment. In this study, we found that when pregnant mice were treated with a high (12.5 mg/g) dose of GABA (orally) during preimplantation, there was a reduction in the number of implantation sites on day 5 of pregnancy. Also, among these unimplanted embryos, most exhibited morphological degeneration and developmental retardation, and only a few of them developed into blastocysts but could not implant into the uterus. Moreover, the expression of uterine receptivity-related factors-LIF, E-cadherin, and HOXA10-were all downregulated, while the number of uterine glands was reduced in the high GABA dose group. Finally, in vitro results demonstrated that GABA (ranging from 10 to 50 µg/µL) markedly inhibited preimplantation embryo development in a dose-response manner. However, this inhibitory effect was not observed when the embryos were pretreated with 40 µΜ 2-hydroxysaclofen, a GABAB antagonist, indicating that GABA exerts its inhibitory effects via its B-type receptor. Our results suggest that exposure to certain GABA concentrations, during early pregnancy, can impair preimplantation embryo development via its B-type receptor, and endometrial receptivity, which greatly disturbs early embryo implantation in mice. These findings could raise concerns about GABA consumption during the early stages of pregnancy.

Korean meditation music suppressed relapse to methamphetamine in rats.

OBJECTIVE: Methamphetamine is a drug abused worldwide. Even though its abuse is a serious problem in many countries, there are few safe and effective therapies to treat addiction. In a previous study, music therapy attenuated relapse to morphine. Based on the study, we investigated whether music therapy suppresses the reinstatement of methamphetamine self-administration behavior. METHODS: Male Sprague-Dawley rats were trained to intravenously self-administer methamphetamine (0.1 mg/kg) using a fixed ratio 1 schedule in a daily 2 h session. Following 3 weeks of training, rats who had established a stable daily intake were subjected to extinction for 1 week. On the next day, priming injection was performed to induce reinstatement. Music therapy was played twice daily during the extinction period and immediately before the test session. In the second experiment, the selective GABAA and GABAB receptor antagonists were treated prior to the last music therapy to investigate a possible neuronal mechanism. Immunofluorescence was performed to immunohistochemically examine the behavioral effects. RESULTS: The meditation music by Young-Dong Kim but not the control music by Jeff Beck reduced active lever responding during the reinstatement session. And the effects of music therapy were blocked by GABA receptors antagonists. Also, immunofluorescence showed the pattern of c-Fos expression in the nucleus accumbens paralleled the behavioral results. CONCLUSION: Results of the present study suggest that meditation music by Young-Dong Kim can be a useful therapy to prevent the reinstatement of methamphetamine addiction during abstinence.

Central, but not systemic, thermoregulatory effects of leptin are impaired in rats with obesity: interactions with GABAB agonist and antagonist.

Leptin is an adipokine that regulates body weight by decreasing appetite and increasing energy expenditure. Besides the effects on food intake, leptin can regulate energy expenditure at least in part by modulating thermogenesis. Many of the effects of leptin are attributable to action in the central nervous system, particularly in the hypothalamus. Common forms of obesity are associated with increased leptin levels and a failure to suppress feeding and mediate weight loss in response to exogenous leptin. This apparent leptin ineffectiveness defines a state of so-called leptin resistance. We examined the effect of leptin on core body temperature in rats with normal weight and diet-induced obesity (DIO), as well as thermoregulatory interactions between leptin and GABAB-agonist and an antagonist. We found that leptin retains the ability to induce hyperthermic effect in rats with DIO. Additionally, temperature responses produced by GABAB agonist and antagonist are altered in a state of obesity and by administration of leptin. We evaluated whether the medial preoptic area of the anterior hypothalamus (MPA) still remains sensitive to leptin action during DIO. Using extracellular recordings of neurons and phospho-signal transducer and the activator of transcription 3 (pSTAT3) immunohistochemistry, we have provided strong evidence that leptin signaling in the MPA is impaired in obese rats. We believe that leptin resistance in the MPA may play a role in the pathogenesis of obesity and obesity-related disease states.

POSSIBLE IMPLEMENTATION OF GABAERGIC AND GLUTAMATERGIC SYSTEMS IN REALIZATION OF ANTIEPILEPTIC EFFECTS OF ACOUSTIC RANGE ELECTRO - MAGNETIC FIELDS.

Seizure is a clinical manifestation of a hyperexcitable neuronal network, in which, the electrical balance underlying the normal neuronal activity is altered pathologically-excitation (Glutamatergic activity) predominates over inhibition (GABAergic activity). Arresting of seizure activity is carried out by restoration of neurotransmitter balance. This process has a direct relation with ion channel permeability in cell and ion transmembrane movement. Low frequency EMS may have a neurostimulating and neuromodulating effect that is based on electromagnetic induction of electric field in the brain. Under the conditions of certain amplitude, frequency and relaxation time low-frequency electromagnetic field (EMF) induces depolarization of separate neurons, and changes the total cortical excitability in case of repeatedly carried out procedures. It was shown that the exposure of acoustic range EMS in GEPRs treated with GABA-A or GABA-B receptors antagonists decreased behavior seizure activity in response to audiogenic stimuli. Injection of Glutamate receptor agonist on background EMS causes seizure activity, but seizure manifestations have less degree compared to non-stimulated rats. Thus, in response to electromagnetic stimulation, the reduction or complete cramping of seizures can be explained by a change in the activity of the neurotransmitter systems.

GABAB receptor regulates proliferation in the high-grade chondrosarcoma cell line OUMS-27 via apoptotic pathways.

BACKGROUND: High-grade chondrosarcoma, which has a high incidence of local recurrence and pulmonary metastasis despite surgical resection, is associated with poor prognosis. Therefore, new and effective adjuvant therapies are urgently required for this disease. Gamma-aminobutyric acid (GABA), which acts as a neurotrophic factor during nervous system development, is related to the proliferation and migration of certain cancer cells. The GABAergic system, which is composed of GABA, the GABA-synthesizing enzyme glutamic acid decarboxylase (GAD), and GABA receptors, has an important function in nerve growth and development of neural crest. Therefore, the GABAergic system may play important functional roles in the proliferation of chondrosarcoma cells, which are derived from neural crest cells. We examined the anti-tumor effects of the GABAergic system on a chondrosarcoma cell line. METHODS: We evaluated the underlying mechanisms of the anti-tumor effects of the GABAergic system, such as the involvement of different signaling pathways, apoptosis, and cell cycle arrest, in the high-grade chondrosarcoma cell line OUMS-27. In addition, we performed whole-cell patch-clamp recordings for Ca2+ currents and evaluated the changes in intracellular Ca2+ concentration via Ca2+ channels, which are related to the GABAB receptor in high-grade chondrosarcoma cells. RESULTS: The GABAB receptor antagonist CGP had anti-tumor effects on high-grade chondrosarcoma cells in a dose-dependent manner. The activities of caspase 3 and caspase 9 were significantly elevated in CGP-treated cells compared to in untreated cells. The activity of caspase 8 did not differ significantly between untreated cells and CGP-treated cells. However, caspase 8 tended to be up-regulated in CGP-treated cells. The GABAB receptor antagonist exhibited anti-tumor effects at the G1/S cell cycle checkpoint and induced apoptosis via dual inhibition of the PI3/Akt/mTOR and MAPK signaling pathways. Furthermore, the changes in intracellular Ca2+ via GABAB receptor-related Ca2+ channels inhibited the proliferation of high-grade chondrosarcoma cells by inducing and modulating apoptotic pathways. CONCLUSIONS: The GABAB receptor antagonist may improve the prognosis of high-grade chondrosarcoma by exerting anti-tumor effects via different signaling pathways, apoptosis, cell cycle arrest, and Ca2+ channels in high-grade chondrosarcoma cells.

Low Cerebral Exposure Cannot Hinder the Neuroprotective Effects of Panax Notoginsenosides.

A bidirectional route of communication between the gastrointestinal tract and the central nervous system, termed the "gut-brain axis," is becoming increasingly relevant to treatment of cerebral damage. Panax Notoginsenoside extract (PNE) is popular for prevention and treatment of cardio-cerebrovascular ischemic diseases although plasma and cerebral exposure levels are extremely low. To date, the mechanisms underlying the neuroprotective effects of PNE remain largely unknown. In the present study, the neuroprotective effects of PNE were systematically studied via investigation of the regulation by PNE of the gastrointestinal microbial community and γ aminobutyric acid (GABA) receptors. The results demonstrated that pretreatment with PNE exerted a remarkable neuroprotective effect on focal cerebral ischemia/reperfusion (I/R) injury in rats, and the efficiency was attenuated in germ-free rats. Pretreatment with PNE could significantly prevent downregulation of Bifidobacterium longum (B.L) caused by I/R surgery, and colonization by B.L could also exert neuroprotective effects. More importantly, both PNE and B.L could upregulate the expression of GABA receptors in the hippocampus of I/R rats, and coadministration of a GABA-B receptor antagonist could significantly attenuate the neuroprotective effects of PNE and B.L. The study above suggests that the neuroprotective effects of PNE may be largely attributable to its regulation of intestinal flora, and oral treatment with B.L was also useful in therapy of ischemia/reperfusion injury (I/R) by upregulating GABA-B receptors.

Acupuncture reduces relapse to cocaine-seeking behavior via activation of GABA neurons in the ventral tegmental area.

There is growing public interest in alternative approaches to addiction treatment and scientific interest in elucidating the neurobiological underpinnings of acupuncture. Our previous studies showed that acupuncture at a specific Shenmen (HT7) points reduced dopamine (DA) release in the nucleus accumbens (NAc) induced by drugs of abuse. The present study was carried out to evaluate the effects of HT7 acupuncture on γ-aminobutyric acid (GABA) neuronal activity in the ventral tegmental area (VTA) and the reinstatement of cocaine-seeking behavior. Using microdialysis and in vivo single-unit electrophysiology, we evaluated the effects of HT7 acupuncture on VTA GABA and NAc DA release and VTA GABA neuronal activity in rats. Using a within-session reinstatement paradigm in rats self-administering cocaine, we evaluated the effects of HT7 stimulation on cocaine-primed reinstatement. Acupuncture at HT7 significantly reduced cocaine suppression of GABA release and GABA neuron firing rates in the VTA. HT7 acupuncture attenuated cocaine-primed reinstatement, which was blocked by VTA infusions of the selective GABAB receptor antagonist 2-hydroxysaclofen. HT7 stimulation significantly decreased acute cocaine-induced DA release in the NAc, which was also blocked by 2-hydroxysaclofen. HT7 acupuncture also attenuated cocaine-induced sensitization of extracellular DA levels in the NAc. Moreover, HT7 acupuncture reduced both locomotor activity and neuronal activation in the NAc induced by acute cocaine in a needle-penetration depth-dependent fashion. These results suggest that acupuncture may suppress cocaine-induced DA release in the NAc and cocaine-seeking behavior through activation of VTA GABA neurons. Acupuncture may be an effective therapy to reduce cocaine relapse by enhancing GABAergic inhibition in the VTA.

Reducing effect of saikosaponin A, an active ingredient of Bupleurum falcatum, on alcohol self-administration in rats: Possible involvement of the GABAB receptor.

Recent studies demonstrated that treatment with saikosaponin A (SSA) - an active ingredient of the medicinal herb, Bupleurum falcatum L. - selectively suppressed, likely via a GABAB receptor-mediated mechanism, intravenous self-administration of morphine and cocaine in rats [Yoon et al., 2012; 2013]. The present study was designed to investigate whether the capacity of SSA to suppress morphine and cocaine self-administration extends to oral alcohol self-administration. To this end, selectively bred Sardinian alcohol-preferring (sP) rats were trained to lever-respond on a Fixed Ratio (FR) 4 (FR4) schedule of reinforcement for alcohol (15%, v/v) in daily 30-min sessions. Once responding had stabilized, rats were tested under the FR4 (measure of alcohol reinforcing properties) and Progressive Ratio (PR; measure of alcohol motivational properties) schedules of reinforcement. The possible involvement of the GABAB receptor system was investigated testing the effect of (a) pretreatment with the GABAB receptor antagonist, SCH50911, and (b) combined treatment with the positive allosteric modulator of the GABAB receptor, GS39783. Treatment with SSA (0, 0.25, 0.5, and 1mg/kg, i.p.) markedly reduced lever-responding for alcohol, amount of self-administered alcohol, and breakpoint for alcohol (defined as the lowest response requirement not achieved in the PR experiment). Pretreatment with 2mg/kg SCH50911 (i.p.) resulted in a partial blockade of the reducing effect of 0.5mg/kg SSA on lever-responding for alcohol and amount of self-administered alcohol. Combination of per se ineffective doses of GS39783 (5mg/kg, i.g.) and SSA (0.1mg/kg, i.p.) reduced lever-responding for alcohol and amount of self-administered alcohol. These results (a) extend to alcohol self-administration the capacity of SSA to suppress morphine and cocaine self-administration in rats and (b) suggest that the GABAB receptor system is likely part of the neural substrate underlying the reducing effect of SSA on alcohol self-administration.

Blockade of in vitro ictogenesis by low-frequency stimulation coincides with increased epileptiform response latency.

Low-frequency stimulation, delivered through transcranial magnetic or deep-brain electrical procedures, reduces seizures in patients with pharmacoresistant epilepsy. A similar control of ictallike discharges is exerted by low-frequency electrical stimulation in rodent brain slices maintained in vitro during convulsant treatment. By employing field and "sharp" intracellular recordings, we analyzed here the effects of stimuli delivered at 0.1 or 1 Hz in the lateral nucleus of the amygdala on ictallike epileptiform discharges induced by the K(+) channel blocker 4-aminopyridine in the perirhinal cortex, in a rat brain slice preparation. We found that 1) ictal events were nominally abolished when the stimulus rate was brought from 0.1 to 1 Hz; 2) this effect was associated with an increased latency of the epileptiform responses recorded in perirhinal cortex following each stimulus; and 3) both changes recovered to control values following arrest of the 1-Hz stimulation protocol. The control of ictal activity by 1-Hz stimulation and the concomitant latency increase were significantly reduced by GABAB receptor antagonism. We propose that this frequency-dependent increase in latency represents a short-lasting, GABAB receptor-dependent adaptive mechanism that contributes to decrease epileptiform synchronization, thus blocking seizures in epileptic patients and animal models.

Acupuncture at HT7 suppresses morphine self-administration at high dose through GABA system.

In the previous study, acupuncture at HT7 has shown to attenuate the self-administration of morphine at a low dose (0.1mg/kg). In this study, it was further investigated whether acupuncture at HT7 could attenuate the morphine self-administration at a high dose (0.5mg/kg). Male Sprague-Dawley rats weighing 270-300g were used. After surgery of catheterization, animals were trained to self-administer morphine solution (0.5mg/kg) using daily 1h session under fixed ratio 1 schedule for 3 weeks. Animals that had shown stable morphine-taking (establish baseline: variation less than 20% of the mean of three consecutive days) were subjected to the acupuncture treatment. Bicuculline and SCH 50911 were used to investigate the possible relation between the effect of acupuncture and the GABA receptor system. Acupuncture at HT7, but not at control acupoint, LI5, suppressed spontaneous morphine-taking behavior significantly. In addition, the effect of acupuncture was blocked by both GABA receptor antagonists. The results of this study suggest that acupuncture at HT7 suppresses morphine-taking behavior through the mediation of GABA receptor system.

Long-lasting hyperpolarization underlies seizure reduction by low frequency deep brain electrical stimulation.

Mesial temporal lobe epilepsy (MTLE) is a common medically refractory neurological disease. Deep brain electrical stimulation (DBS) of grey matter has been used for MTLE with limited success. However, stimulation of a white matter tract connecting the hippocampi, the ventral hippocampal commissure (VHC), with low frequencies that simulate interictal discharges has shown promising results, with seizure reduction greater than 98% in bilateral hippocampi during stimulation and greater than 50% seizure reduction in bilateral hippocampi after treatment. A major hurdle to the implementation and optimization of this treatment is that the mechanisms of seizure reduction by low frequency electrical stimulation (LFS) are not known. The goal of this study is to understand how commissural fibre tract stimulation reduces bilateral hippocampal epileptic activity in an in vitro slice preparation containing bilateral hippocampi connected by the VHC. It is our hypothesis that electrical stimuli induce hyperpolarization lasting hundreds of milliseconds following each pulse which reduces spontaneous epileptic activity during each inter-stimulus interval (ISI). Stimulus-induced long-lasting-hyperpolarization (LLH) can be mediated by GABA(B) inhibitory post-synaptic potentials (IPSPs) or slow after-hyperpolarization (sAHP). To test the role of LLH in effective bilateral seizure reduction by fibre tract stimulation, we measured stimulus-induced hyperpolarization during LFS of the VHC using electrophysiology techniques. Antagonism of the GABA(B) IPSP and/or sAHP diminished stimulus-induced hyperpolarization concurrently with LFS efficacy (greater than 50% reduction). Blocking both the GABA(B) IPSP and sAHP simultaneously eliminated the effect of electrical stimulation on seizure reduction entirely. These data show that LFS of the VHC is an effective protocol for bilateral hippocampal seizure reduction and that its efficacy relies on the induction of long-lasting hyperpolarization mediated through GABA(B) IPSPs and sAHP. Based on this study, optimization of the timing of LFS and LFS-induced-LLH may lead to improved outcomes from DBS treatments for human epilepsy.

Involvement of hypothalamic periventricular GABAergic nerves in the central pressor response to clonidine in freely-moving conscious rats.

Microinjection of the α(2)-adrenoceptor agonist clonidine into the hypothalamic periventricular nuclei (PVN) induces the pressor response associated with bradycardia in freely-moving conscious rats. This study investigated the involvement of γ-aminobutyric acid nerves (GABAergic nerves) and glutamatergic nerves in the cardiovascular response to microinjection of clonidine in the PVN. Male Wistar rats were chronically implanted with a microinjection cannula into the PVN and an arterial catheter into the abdominal aorta through the femoral artery. Blood pressure and heart rate were measured under a conscious unrestrained state. PVN injection of clonidine induced a dose-dependent pressor response concomitant with bradycardia. PVN pretreatment with GABA, muscimol (GABA(A)-receptor agonist), or bicuculline (GABA(A)-receptor antagonist) significantly inhibited the pressor response to PVN-injected clonidine without affecting bradycardia. PVN pretreatment with baclofen (GABA(B)-receptor agonist), 2-hydroxysaclofen (GABA(B)-receptor antagonist), or kynurenic acid (non-selective NMDA-type glutamate-receptor and ionotropic glutamate-receptor antagonist) did not affect the pressor response to PVN-injected clonidine. These results suggest that clonidine induces a pressor response by stimulating the presynaptic α(2)-adrenoceptor of GABAergic nerves in the PVN, thereby inhibiting GABAergic nerve activity.

Effect of acupuncture on naloxone-precipitated withdrawal syndrome in morphine-experienced rats: the mediation of GABA receptors.

Repeated morphine administration increases extracellular dopamine levels in the nucleus accumbens, which results in behavioral sensitization that can be suppressed by acupuncture at Shenmen (HT7) points. The present study was conducted to investigate the effects of acupuncture at HT7 on morphine withdrawal syndrome as well as to explore the role of GABA receptors in mediating the effects of HT7 acupuncture. We induced morphine withdrawal by injecting naloxone to rats that self-administer morphine and evaluated the effects of acupuncture and/or GABA receptor antagonists on their withdrawal symptoms. Acupuncture at HT7, but not at the control point LI5, significantly decreased symptoms of morphine withdrawal. HT7 inhibition of the withdrawal syndrome was blocked by pretreatment with either the GABA(A) receptor antagonist bicuculline or the GABA(B) antagonist SCH 50911. These findings suggest that the effects of acupuncture on suppression of morphine withdrawal syndrome are mediated, at least in part, through GABA receptors.

The GABA B receptor agonist CGP44532 and the positive modulator GS39783 reverse some behavioural changes related to positive syndromes of psychosis in mice.

BACKGROUND AND PURPOSE: An important role of GABAergic neurotransmission in schizophrenia was proposed a long time ago, but there is limited data to support this hypothesis. In the present study we decided to investigate GABA(B) receptor ligands in animal models predictive for the antipsychotic activity of drugs. The GABA(B) receptor antagonists CGP51176 and CGP36742, agonist CGP44532 and positive allosteric modulator GS39783 were studied. EXPERIMENTAL APPROACH: The effects of all ligands were investigated in MK-801- and amphetamine-induced hyperactivity tests. The anti-hallucinogenic-like effect of the compounds was screened in the model of head twitches induced by (±)1-(2.5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI). Furthermore, the effect of GS39783 and CGP44532 on DOI-induced frequency of spontaneous excitatory postsynaptic currents (EPSCs) in slices from mouse brain frontal cortices was investigated. The anti-cataleptic properties of the compounds were also assessed. KEY RESULTS: The GABA(B) receptor activators CGP44532 and GS39783 exhibited antipsychotic-like effects both in the MK-801- and amphetamine-induced hyperactivity tests, as well as in the head-twitch model in mice. Such effects were not observed for the GABA(B) receptor antagonists. DOI-induced increased frequency of spontaneous EPSCs was also decreased by the compounds. Moreover, CGP44532 and GS39783 inhibited haloperidol-induced catalepsy and EPSCs. CONCLUSION AND IMPLICATIONS: These data suggest that selective GABA(B) receptor activators may be useful in the treatment of psychosis.