Glycinergic Activity Research Articles

Loss of glycine receptors in the nucleus accumbens and ethanol reward in an Alzheimer´s Disease mouse model

Alterations in cognitive and non-cognitive cerebral functions characterize Alzheimer's disease (AD). Cortical and hippocampal impairments related to extracellular accumulation of Aβ in AD animal models have been extensively investigated. However, recent reports have also implicated intracellular Aβ in limbic regions, such as the nucleus accumbens (nAc). Accumbal neurons express high levels of inhibitory glycine receptors (GlyRs) that are allosterically modulated by ethanol and have a role in controlling its intake. In the present study, we investigated how GlyRs in the 2xTg mice (AD model) affect nAc functions and ethanol intake behavior. Using transgenic and control aged-matched litter mates, we found that the GlyRα2 subunit was significantly decreased in AD mice (6-month-old). We also examined intracellular calcium dynamics using the fluorescent calcium protein reporter GCaMP in slice photometry. We also found that the calcium signal mediated by GlyRs, but not GABAAR, was also reduced in AD neurons. Additionally, ethanol potentiation was significantly decreased in accumbal neurons in the AD mice. Finally, we performed drinking in the dark (DID) experiments and found that 2xTg mice consumed less ethanol on the last day of DID, in agreement with a lower blood ethanol concentration. 2xTg mice also showed lower sucrose consumption, indicating that overall food reward was altered. In conclusion, the data support the role of GlyRs in nAc neuron excitability and a decreased glycinergic activity in the 2xTg mice that might lead to impairment in reward processing at an early stage of the disease.

Novel Phenylene Lipids That Are Positive Allosteric Modulators of Glycine Receptors and Inhibitors of Glycine Transporter 2.

Chronic pain is a complex condition that remains resistant to current therapeutics. We previously synthesized a series of N-acyl amino acids (NAAAs) that inhibit the glycine transporter, GlyT2, some of which are also positive allosteric modulators of glycine receptors (GlyRs). In this study, we have synthesized a library of NAAAs that contain a phenylene ring within the acyl tail with the objective of improving efficacy at both GlyT2 and GlyRs and also identifying compounds that are efficacious as dual-acting modulators to enhance glycine neurotransmission. The most efficacious positive allosteric modulator of GlyRs was 2-[8-(2-octylphenyl)octanoylamino]acetic acid (8-8 OPGly) which potentiates the EC5 for glycine activation of GlyRα1 by 1500% with an EC50 of 664 nM. Phenylene-containing NAAAs with a lysine headgroup were the most potent inhibitors of GlyT2 with (2S)-6-amino-2-[8-(3-octylphenyl)octanoylamino]hexanoic acid (8-8 MPLys) inhibiting GlyT2 with an IC50 of 32 nM. The optimal modulator across both proteins was (2S)-6-amino-2-[8-(2-octylphenyl)octanoylamino]hexanoic acid (8-8 OPLys), which inhibits GlyT2 with an IC50 of 192 nM and potentiates GlyRs by up to 335% at 1 μM. When tested in a dual GlyT2/GlyRα1 expression system, 8-8 OPLys caused the greatest reductions in the EC50 for glycine. This suggests that the synergistic effects of a dual-acting modulator cause greater enhancements in glycinergic activity compared to single-target modulators and may provide an alternate approach to the development of new non-opioid analgesics for the treatment of chronic pain.

Glycine neurotransmission: Its role in development

The accurate function of the central nervous system (CNS) depends of the consonance of multiple genetic programs and external signals during the ontogenesis. A variety of molecules including neurotransmitters, have been implied in the regulation of proliferation, survival, and cell-fate of neurons and glial cells. Among these, neurotransmitters may play a central role since functional ligand-gated ionic channel receptors have been described before the establishment of synapses. This review argues on the function of glycine during development, and show evidence indicating it regulates morphogenetic events by means of their transporters and receptors, emphasizing the role of glycinergic activity in the balance of excitatory and inhibitory signals during development. Understanding the mechanisms involved in these processes would help us to know the etiology of cognitive dysfunctions and lead to improve brain repair strategies.

Sarcophine and (7S, 8R)-dihydroxydeepoxysarcophine from the Red Sea soft coral Sarcophyton glaucum as in vitro and in vivo modulators of glycine receptors

The inhibitory glycine receptor (GlyR) is a key mediator of synaptic signalling in spinal cord, brain stem, and higher centres of the central nervous system. We examined the glycinergic activity of sarcophine (SN), a marine terpenoid known for its various biological activities, and its trans-diol derivative (7S, 8R)-dihydroxy-deepoxysarcophine (DSN). SN was isolated from the Red Sea soft coral Sacrophyton glaucum, DSN was semisynthesized by hydrolysis of the epoxide ring. In cytotoxicity tests against HEK293 cells, SN and DSN had LD50 values of 29.3 ± 3.0 mM and 123.5 ± 13.0 mM, respectively. Both compounds were tested against recombinant human α1 glycine receptors in HEK293 cells using whole-cell recording techniques. Both, SN and DSN were shown for the first time to be inhibitors of recombinant glycine receptors, with KIvalues of 2.1 ± 0.3 μM for SN, and 109 ± 9 μM for DSN. Receptor inhibition was also studied in vivo in a mouse model of strychnine toxicity. Surprisingly, in mouse experiments strychnine inhibition was not augmented by either terpenoid. While DSN had no significant effect on strychnine toxicity, SN even delayed strychnine effects. This could be accounted for by assuming that strychnine and sarcophine derivatives compete for the same binding site on the receptor, so the less toxic sarcophine can prevent strychnine from binding. The combination of modulatory activity and low level of toxicity makes sarcophines attractive structures for novel glycinergic drugs.

Glycine Promotes the Survival of a Subpopulation of Neural Stem Cells.

Glycine is mainly known as an inhibitory neurotransmitter in adult mature neurons, regulating neuronal network activity in the central nervous system. In contrast, during embryogenesis glycine can act as an excitatory neurotransmitter and generates the first electrical signal in immature neurons. The roles and functional significance of this excitatory glycinergic activity during neurodevelopment are still unclear. Using the zebrafish embryo as a model, we previously showed that glycine regulates proliferation and differentiation of neural stem cells (NSCs) to interneurons. Moreover, we identified that glycine signaling in NSCs is associated with several common developmental pathways and surprisingly also the p53-related apoptosis. Here we investigated how glycine signaling regulates NSC survival. First, we showed by two approaches, acridine orange staining and active caspase 3 immunostaining that defects in glycine signaling induce an early and transient cell death, which was suppressed by knockdown of p53. Then, we developed an NSC transplantation strategy to directly assess NSC-autonomous development upon perturbing glycine signaling. In vivo time-lapse imaging showed that disruption of glycine signaling disturbed the normal NSC interkinetic nuclear migration, leading to cell cycle arrest and apoptosis. Finally, we analyzed two main subpopulations of NSCs, expressing either nestin or GFAP, by in situ labeling and in transgenic lines expressing GFP in either population. We found that disruption of glycine signaling induced a drastic and selective loss of nestin-positive (nestin+) NSCs, which was only partially rescued upon p53 knockdown. Taken together, our findings support a role of glycine signaling in promoting survival of the nestin+ NSC subpopulation early during development.

Evaluation and Comparison of Anti-Epileptic Activity of Methanolic Extracts of Seeds of Cuminum Cyminum L. and Centratherum Anthelminticum L. in Seizure Model

Objective: Evaluation and comparison of anti-epileptic activity of methanolic extracts of seeds of Cuminum cyminum L. and Centratherum anthelminticum L. in strychnine-induced seizure model.
 Methods: The study was conducted in the Department of Pharmacology, University of Karachi. After 15 days of dosing, experiment was performed. A total of 40 healthy mice were selected from animal house of the Pharmacology department, University of Karachi, and were equally divided into four groups. Group 01 was on DMSO (dimethyl sulphoxide) organic solvent as control; Group 02 was on methanolic extract of Cuminum cyminum L. (500 mg/kg); Group 03 was on methanolic extract of Centratherum anthelminticum L. (200 mg/kg) and Group 04 was on reference drug diazepam (3 mg/kg). On the day of experiment, after giving doses of DMSO, methanolic extract of Cuminum cyminum L., methanolic extract of Centratherum anthelminticum L. and diazepam, respectively to all groups, strychnine was administered and animals were observed for latency period and duration of Mortality rate was also evaluated.
 Results: Group 02 and 03, receiving methanolic extracts of Cuminum cyminum L. and Centratherum anthelminticum L. showed significant seizure protection as observed by delayed seizure onset (p <0.001) and decreased total duration of convulsions in groups 02 and 03 (p£ 0.01, p£ 0.05) respectively. The mortality rate also decreased significantly (p £ 0.05) in comparison with group 01.
 Conclusion: Anticonvulsant activity of methanolic extracts of Cuminum cyminum L. and Centratherum anthelminticum L. could be due to their antioxidant activity and further enhanced by potentiation of GABAergic and glycinergic activity. Further studies are required to confirm exact mechanism of action.

Second Messengers in the Presynaptic Regulation of Glycinergic Synapses in Frog Motoneurons

The possible pathways mediating the inhibitory influences of two types of presynaptic metabotropic receptors – group III metabotropic glutamate receptors (mGluRIII) and GABAB receptors (GABABR) – on miniature glycinergic activity in motoneurons in the isolated spinal cord of the frog Rana ridibunda were studied. The glycinergic component is dominant in the inhibitory miniature activity of motoneurons [3]. Baclofen and LAP4, which are selective agonists of GABABR and mGluRIII, inhibited miniature glycinergic activity, decreasing frequency by 48.0 ± 5.6 (n = 8) and 48.5 ± 8.6% (n = 5), respectively. The amplitude of miniature potentials did not change significantly, which is evidence for a presynaptic mechanism of modulation of synaptic transmission. Studies of the stages in the mediation of the influences of the metabotropic receptors studied here on the process of glycine release from presynaptic terminals were performed by applying their selective agonists in conditions of previous selective blockade of the components mediating modulation of glycinergic synaptic transmission. The effects of mGluRIII were shown to be mediated tghe feedback via adenylate cyclase (AC), the next stage being mediated by protein kinase A (PKA), as their selective blockade virtually eliminated by effect of activation of mGluRIII. The effects of GABABR on miniature glycinergic activity did not involve AC activity, as its blocker SQ22536 did not alter the inhibitory effect of baclofen. The effects of GABABR were mediated by negative feedback via phospholipase C (PLC), as its prior exclusion (inhibitor U73122) prevented the effect of application of baclofen. However, the next possible stage in this pathway – protein kinase C (PKC) – did not have a role in mediating the influences of GABABR, as the PKC blocker (GF109302X) had no influences on the effect of baclofen. The common component in mediating the effects of mGluRIII and GABABR on miniature glycinergic activity may consist of inositol triphosphate receptors (IP3R), which regulate calcium release from intraterminal depots and, according to published data, are linked with both PKA or cAMP and with PLC. In our experiments, 2-APB, an IP3R antagonist, decreased miniature glycinergic activity frequency by 26 ± 8% (n = 7) and prevented the inhibitory effects of application of both mGluRIII and GABABR agonists. This stage of mediation of the influences of mGluRIII and GABABR appears to involve collision and interaction between these (crosstalk), as demonstrated by our previous studies [2].

D-cycloserine improves synaptic transmission in an animal mode of Rett syndrome

Rett syndrome (RTT), a leading cause of intellectual disability in girls, is predominantly caused by mutations in the X-linked gene MECP2. Disruption of Mecp2 in mice recapitulates major features of RTT, including neurobehavioral abnormalities, which can be reversed by re-expression of normal Mecp2. Thus, there is reason to believe that RTT could be amenable to therapeutic intervention throughout the lifespan of patients after the onset of symptoms. A common feature underlying neuropsychiatric disorders, including RTT, is altered synaptic function in the brain. Here, we show that Mecp2tm1.1Jae/y mice display lower presynaptic function as assessed by paired pulse ratio, as well as decreased long term potentiation (LTP) at hippocampal Schaffer–collateral-CA1 synapses. Treatment of Mecp2tm1.1Jae/y mice with D-cycloserine (DCS), an FDA-approved analog of the amino acid D-alanine with antibiotic and glycinergic activity, corrected the presynaptic but not LTP deficit without affecting deficient hippocampal BDNF levels. DCS treatment did, however, partially restore lower BDNF levels in the brain stem and striatum. Thus, treatment with DCS may mitigate the severity of some of the neurobehavioral symptoms experienced by patients with Rett syndrome.

Neurotransmission to parasympathetic cardiac vagal neurons in the brain stem is altered with left ventricular hypertrophy-induced heart failure.

Hypertension, cardiac hypertrophy, and heart failure (HF) are widespread and debilitating cardiovascular diseases that affect nearly 23 million people worldwide. A distinctive hallmark of these cardiovascular diseases is autonomic imbalance, with increased sympathetic activity and decreased parasympathetic vagal tone. Recent device-based approaches, such as implantable vagal stimulators that stimulate a multitude of visceral sensory and motor fibers in the vagus nerve, are being evaluated as new therapeutic approaches for these and other diseases. However, little is known about how parasympathetic activity to the heart is altered with these diseases, and this lack of knowledge is an obstacle in the goal of devising selective interventions that can target and selectively restore parasympathetic activity to the heart. To identify the changes that occur within the brain stem to diminish the parasympathetic cardiac activity, left ventricular hypertrophy was elicited in rats by aortic pressure overload using a transaortic constriction approach. Cardiac vagal neurons (CVNs) in the brain stem that generate parasympathetic activity to the heart were identified with a retrograde tracer and studied using patch-clamp electrophysiological recordings in vitro. Animals with left cardiac hypertrophy had diminished excitation of CVNs, which was mediated both by an augmented frequency of spontaneous inhibitory GABAergic neurotransmission (with no alteration of inhibitory glycinergic activity) as well as a diminished amplitude and frequency of excitatory neurotransmission to CVNs. Opportunities to alter these network pathways and neurotransmitter receptors provide future targets of intervention in the goal to restore parasympathetic activity and autonomic balance to the heart in cardiac hypertrophy and other cardiovascular diseases.

C-fiber activity-dependent maturation of glycinergic inhibition in the spinal dorsal horn of the postnatal rat.

Sensory circuits are shaped by experience in early postnatal life and in many brain areas late maturation of inhibition drives activity-dependent development. In the newborn spinal dorsal horn, activity is dominated by inputs from low threshold A fibers, whereas nociceptive C-fiber inputs mature gradually over the first postnatal weeks. How this changing afferent input influences the maturation of dorsal horn inhibition is not known. We show an absence of functional glycinergic inhibition in newborn dorsal horn circuits: Dorsal horn receptive fields and afferent-evoked excitation are initially facilitated by glycinergic activity due, at least in part, to glycinergic disinhibition of GAD67 cells. Glycinergic inhibitory control emerges in the second postnatal week, coinciding with an expression switch from neonatal α(2) homomeric to predominantly mature α(1)/β glycine receptors (GlyRs). We further show that the onset of glycinergic inhibition depends upon the maturation of C-fiber inputs to the dorsal horn: selective block of afferent C fibers in postnatal week 2, using perisciatic injections of the cationic anesthetic QX-314, lidocaine, and capsaicin, delays the maturation of both GlyR subunits and glycinergic inhibition, maintaining dorsal neurons in a neonatal state, where tactile responses are facilitated, rather than inhibited, by glycinergic network activity. Thus, glycine may serve to facilitate tactile A-fiber-mediated information and enhance activity-dependent synaptic strengthening in the immature dorsal horn. This period ceases in the second postnatal week with the maturation of C-fiber spinal input, which triggers postsynaptic changes leading to glycinergic inhibition and only then is balanced excitation and inhibition achieved in dorsal horn sensory circuits.

Oestradiol Rapidly Enhances Spontaneous Glycinergic Synaptic Inhibition of Hypoglossal Motoneurones

Whereas oestradiol is well-known to facilitate excitatory glutamatergic synaptic transmission, its effects on fast inhibitory neurotransmission are not as well established. Possible acute modulation of the spontaneous glycinergic synaptic activity by oestradiol was investigated in voltage-clamped hypoglossal motoneurones by whole-cell patch-clamp recording in rat brainstem slices. The spontaneous glycinergic synaptic activity was continuously recorded in each neurone under control conditions, during 12-20 min of perfusion with 17beta-oestradiol and during washing. When oestradiol was diluted in ethanol, the control solution contained the same amount of ethanol. At 100 nM, oestradiol markedly increased the frequency of the total spontaneous glycinergic activity. Similar experiments were performed after blockade of action potentials by tetrodotoxin, aiming to isolate miniature glycinergic synaptic currents. Oestradiol increased the frequency of glycinergic miniatures in most slices, in some cases within less than 1 min. In some slices, oestradiol also favoured the occurrence of glycinergic miniatures of large amplitude. These effects were slowly reversible during washing. At 1 nm, oestradiol still increased the frequency of glycinergic miniatures. The results were confirmed in the absence of ethanol by using water-soluble cyclodextrin-encapsulated oestradiol. In these experiments, the control solution contained the same amount of (2-hydroxypropyl)-beta-cyclodextrin as the oestradiol-containing solution. In addition, prolonged control recordings were performed without applying oestradiol to check the stability of the glycinergic synaptic activity during prolonged whole-cell recordings. The results show, for the first time, that, within a few minutes, oestradiol can enhance the spontaneous synaptic release of a major inhibitory transmitter, glycine.

NO Differentially Regulates Neurotransmission to Premotor Cardiac Vagal Neurons in the Nucleus Ambiguus

NO is involved in the neural control of heart rate, and NO synthase expressing neurons and terminals have been localized in the nucleus ambiguus where parasympathetic cardiac vagal preganglionic neurons are located; however, little is known about the mechanisms by which NO alters the activity of premotor cardiac vagal neurons. This study examines whether the NO donor sodium nitroprusside ([SNP] 100 micromol/L) and precursor, l-arginine (10 mmol/L), modulate excitatory and inhibitory synaptic neurotransmission to cardiac vagal preganglionic neurons. Glutamatergic, GABAergic, and glycinergic activity to cardiac vagal neurons was examined using whole-cell patch-clamp recordings in an in vitro brain slice preparation in rats. Both SNP, as well as l-arginine, increased the frequency of GABAergic neurotransmission to cardiac vagal preganglionic neurons but decreased the amplitude of GABAergic inhibitory postsynaptic currents. In contrast, both l-arginine and SNP inhibited the frequency of glutamatergic and glycinergic synaptic events in cardiac vagal preganglionic neurons. SNP and l-arginine also decreased glycinergic inhibitory postsynaptic current amplitude, and this response persisted in the presence of tetrodotoxin. Inclusion of the NO synthase inhibitor 7-nitroindazole (100 mumol/L) prevented the l-arginine-evoked responses. These results demonstrate that NO differentially regulates excitatory and inhibitory neurotransmission, facilitating GABAergic and diminishing glutamatergic and glycinergic neurotransmission to cardiac vagal neurons.

Emergence of Action Potential Generation and Synaptic Transmission in Vestibular Nucleus Neurons

Principal cells of the chick tangential nucleus are vestibular nucleus neurons in the hindbrain. Although detailed information is available on the morphogenesis of principal cells and synaptogenesis of primary vestibular fibers, this is the first study of their early functional development, when vestibular terminals emerge at embryonic days 10 and 13 (E10 and E13). At E10, 60% of principal cells generated spikes on depolarization, whereas 50% exhibited excitatory postsynaptic currents (EPSCs) on vestibular-nerve stimulation. The frequency was 0.2 Hz for glutamatergic spontaneous EPSCs (sEPSCs) at -60 mV, and 0.6 Hz for spontaneous inhibitory postsynaptic current (sIPSC) at +10 mV and completely GABAergic. All of these synaptic events were TTX-insensitive, miniature events. At E13, 50% of principal cells generated spikes on depolarization and 82% exhibited EPSCs on vestibular-nerve stimulation. The frequency was 0.7 Hz for sEPSCs at -60 mV, and 0.8 Hz for sIPSCs at +10 mV. Most principal cells had sIPSCs composed of both GABAergic (75%) and glycinergic (25%) events, but a few cells had only GABAergic sIPSCs. TTX decreased the frequency of EPSCs by 12%, and the IPSCs by 17%. In summary, at E10, some principal cells generated immature spikes on depolarization and EPSCs on vestibular-nerve stimulation. At E10, GABAergic events predominated, AMPA events had low frequencies, and glycinergic activity was absent. By E13, glycinergic events first appeared. This data were compared systematically to that obtained from the late-term embryo and hatchling to reveal the long-term sequence of changes in synaptic events and excitability and offer a broader understanding of how the vestibular system is assembled during development.

Fentanyl induced decrease in respiratory sinus arrhythmia may be reversed by activation of 5‐HT4a receptors

Opiate exposure induces a centrally mediated respiratory inhibition, bradycardia, and a reduction in respiratory sinus arrhythmia (RSA). Previous work has shown that RSA is primarily mediated by an increase in both inhibitory GABAergic and glycinergic activity to cardiac vagal neurons (CVNs) during inspiration, and the mu-opioid receptor agonist, fentanyl inhibits both inspiratory neurons and GABAergic neurotransmission to CVNs. 5-HT4a receptor agonists have been shown to reverse fentanyl-induced respiratory depression. This study examines whether the 5-HT4a agonist, BIMU-8, reverses the fentanyl induced decrease in inspiratory-evoked GABA neurotransmission to CVNs. CVNs were identified by the presence of a retrograde fluorescent tracer and inspiratory evoked GABAergic synaptic currents were examined using whole-cell patch-clamp techniques in an in-vitro preparation that spontaneously generates rhythmic respiratory activity. Inspiratory activity was recorded by monitoring the motor neuron population activity from hypoglossal nerve roolets. BIMU-8 was administrated at a dose of 10microM alone or 10 minutes after fentanyl application. Application of BIMU-8 alone increased inspiratory burst frequency by 50%. Co-application of BIMU-8 with fentanyl prevented the opioid induced decrease in inspiratory-evoked GABAergic neurotransmission to cardiac parasympathetic neurons. This work demonstrates 5-HT4a specific receptor agonists overcome mu-opioid receptor induced respiration and RSA depression and may provide a potential therapeutic target for opiate treatment. Supported by NIH grants HL59895 and 72006 to DM.

Ketamine Inhibits Inspiratory-evoked γ-Aminobutyric Acid and Glycine Neurotransmission to Cardiac Vagal Neurons in the Nucleus Ambiguus

Ketamine can be used for perioperative pain management as well as a dissociative anesthetic agent in emergency situations. However, ketamine can induce both cardiovascular and respiratory depression, especially in pediatric patients. Although ketamine has usually been regarded as sympathoexcitatory, recent work has demonstrated that ketamine has important actions on parasympathetic cardiac vagal efferent activity. The current study tests the hypothesis that ketamine, at clinical relevant concentrations, alters central cardiorespiratory interactions in the brainstem and, in particular, the inspiration-evoked increase in gamma-aminobutyric acid-mediated and glycinergic neurotransmission to parasympathetic cardiac efferent neurons. Cardiac vagal neurons were identified by the presence of a retrograde fluorescent tracer. Respiratory evoked gamma-aminobutyric acid-mediated and glycinergic synaptic currents were recorded in cardiac vagal neurons using whole cell patch clamp techniques while spontaneous rhythmic respiratory activity was recorded simultaneously. : Ketamine, at concentrations from 0.1 to 10 microM, evoked a concentration-dependent inhibition of inspiratory burst frequency. Inspiration-evoked gamma-aminobutyric acid-mediated neurotransmission to cardiac vagal neurons was inhibited at ketamine concentrations of 0.5 and 1 microM. The increase in glycinergic activity to cardiac vagal neurons during inspiration was also inhibited at ketamine concentrations of 0.5 and 1 microM. At clinically relevant concentrations (0.5 and 1 microM), ketamine alters central respiratory activity and diminishes both inspiration-evoked gamma-aminobutyric acid-mediated and glycinergic neurotransmission to parasympathetic cardiac efferent neurons. This reduction in inhibitory neurotransmission to cardiac vagal neurons is likely responsible for the compromised respiratory sinus arrhythmia that occurs with ketamine anesthesia.

The ZebrafishshockedGene Encodes a Glycine Transporter and Is Essential for the Function of Early Neural Circuits in the CNS

shocked (sho) is a zebrafish mutation that causes motor deficits attributable to CNS defects during the first2dof development. Mutant embryos display reduced spontaneous coiling of the trunk, diminished escape responses when touched, and an absence of swimming. A missense mutation in the slc6a9 gene that encodes a glycine transporter (GlyT1) was identified as the cause of the sho phenotype. Antisense knock-down of GlyT1 in wild-type embryos phenocopies sho, and injection of wild-type GlyT1 mRNA into mutants rescues them. A comparison of glycine-evoked inward currents in Xenopus oocytes expressing either the wild-type or mutant protein found that the missense mutation results in a nonfunctional transporter. glyt1 and the related glyt2 mRNAs are expressed in the hindbrain and spinal cord in nonoverlapping patterns. The fact that these regions are known to be required for generation of early locomotory behaviors suggests that the regulation of extracellular glycine levels in the CNS is important for proper function of neural networks. Furthermore, physiological analysis after manipulation of glycinergic activity in wild-type and sho embryos suggests that the mutant phenotype is attributable to elevated extracellular glycine within the CNS.

The Estrogen Receptor Modulator Tamoxifen Enhances Spontaneous Glycinergic Synaptic Inhibition of Hypoglossal Motoneurons

Tamoxifen (Tam), a widely used anticancer agent, is now also used for healthy women with risk of breast cancer. Furthermore, it is the prototype of the selective estrogen receptor modulator family, with promise for neuroprotection. However, possible effects on neurotransmission have been little explored. Recently, Tam was shown to potentiate chloride responses to low concentrations of exogenous glycine in cultured spinal neurons from rat embryo. The present study investigates the possible modulation by Tam of the spontaneous synaptic glycinergic activity recorded from voltage-clamped hypoglossal motoneurons, using the whole-cell patch-clamp technique in brainstem slices from juvenile rat. Miniature currents were isolated with tetrodotoxin. Tam increased the mean amplitude of glycinergic miniature currents, by 68-79% at 2 microM (in nine of 10 cells) and by 47% at 0.5 microM (in four of nine cells). Furthermore, Tam markedly increased the frequency of glycinergic miniatures, by a factor reaching 15 in some neurons, even in the presence of the Ca2+ channel blocker Cd2+. Tam also increased the frequency of the total spontaneous glycinergic activity without tetrodotoxin. The increase in miniature amplitude is consistent with the increase in postsynaptic glycine receptor sensitivity previously reported. The increase in frequency indicates an additional presynaptic effect. Addition of exogenous glycine could also increase the frequency of glycinergic miniatures. Thus, one of the presynaptic effects of Tam might be potentiation of the basal activity of presynaptic glycine receptors facilitating glycine release. Possible risks related to modulation of glycinergic neurotransmission by Tam should be considered when recommending its use in healthy individuals.

Spontaneous synaptic activity in chick vestibular nucleus neurons during the perinatal period

The principal cells of the chick tangential nucleus are second-order vestibular neurons involved in the vestibuloocular and vestibulocollic reflexes. The spontaneous synaptic activity of morphologically identified principal cells was characterized in brain slices from 1-day-old hatchlings (H1) using whole-cell voltage-clamp recordings and Cs-gluconate pipet solution. The frequency was 1.45 Hz for spontaneous excitatory postsynaptic currents (sEPSCs) and 1.47 Hz for spontaneous inhibitory postsynaptic currents (sIPSCs). Using specific neurotransmitter receptor antagonists, all of the sEPSCs were identified as AMPA receptor-mediated events, whereas 56% of the sIPSCs were glycine and 44% were GABA A receptor-mediated events. On exposure to TTX, the frequency of EPSCs decreased by 68%, while the frequency of IPSCs decreased by 33%, indicating greater EPSC dependency on presynaptic action potentials. These data on spontaneous synaptic activity at H1 were compared with those obtained in previous studies of 16-day old embryos (E16). After birth, the spontaneous synaptic activity exhibited increased EPSC frequency, increased ratio for excitatory to inhibitory events, increased percentage of TTX-dependent EPSCs, and faster kinetics. In addition, the ratio for glycine/GABA receptor-mediated events increased significantly. Altogether, these data indicate that at hatching spontaneous synaptic activity of vestibular nucleus neurons in brain slices of the chick tangential nucleus undergoes appreciable changes, with increased frequency of EPSCs and glycinergic activity playing more important roles compared with the late-term chick embryo when GABAergic activity prevailed. The definition of this developmental pattern of synaptic activity in vestibular nucleus neurons should contribute to understanding how vestibular reflex activity is established in the hatchling chick.

The therapeutic potential of glycine transporter-1 inhibitors

While current antipsychotic medications are often efficacious for the positive symptoms of schizophrenia, there remains a critical need for compounds with improved tolerability and efficacy for the negative symptoms and cognitive dysfunction associated with this disease. There is a growing body of evidence suggesting that the potentiation of N-methyl-D-aspartate (NMDA) receptor function may be a useful approach for the treatment of schizophrenia. One proposed strategy for this potentiation is to increase synaptic levels of the neurotransmitter glycine by blocking the glycine transporter-1. Since glycine acts as a required co-agonist for the NMDA receptor complex; this approach allows an increase in the effectiveness of normal glutamatergic signalling at the NMDA receptor complex. Recent preclinical research, focused on the development and testing of novel glycine transporter-1 inhibitors, suggests that this approach may be feasible. Converging clinical evidence suggesting therapeutic efficacy following the potentiation of glycinergic activity further supports this approach. Clinical studies with novel glycine re-uptake inhibitors will provide critical information regarding the therapeutic utility and tolerability of this treatment for schizophrenia and other disorders associated with NMDA receptor hypofunction.

Effects of Taurine Deficiency and Chronic Methanol Administration on Rat Retina, Optic Nerve and Brain Amino Acids and Monoamines

A chronic methanol (MeOH) intoxication scheme (2 g/kg/day ip for 2 weeks) was carried out in Sprague-Dawley rats, previously depleted of folates with methotrexate (MTX). β-Alanine (β-Ala), 5%, was also administered to some animals in the drinking water. Amino acids were determined in plasma, retina, optic nerve, hippocampus and posterior cortex by HPLC with fluorescence detection and monoamines in retina, hippocampus and posterior cortex by electrochemical detection. β-Ala administration reduced taurine (Tau) levels in plasma, hippocampus and posterior cortex, but not in retina and optic nerve. Aspartate (Asp) concentration in the optic nerve was increased in MTX-MeOH treated animals, and the administration of β-Ala did not modify this elevation. The association of β-Ala with MTX-MeOH produced an increase of threonine, and a decrease of 5-hydroxytryptamine (5-HT) in the retina without modifying 5-hydroxyindoleacetic acid, whereas in the hippocampus an elevation of asparagine was observed. We conclude that, in the retina, β-Ala in combination with MTX-MeOH increased serotonin and decreased dopamine (DA) turnover rate, and resulted in changes in the amino acid balance, that could affect glycinergic activity. On the other hand, in the hippocampus, Asp metabolism could be affected by Tau depletion with β-Ala.