Structure, diversity and synaptic localization of inhibitory glycine receptors

doi:10.1016/0928-4257(94)90087-6

Journal of Physiology-Paris

Volume 88, Issue 4, 1994, Pages 243-248

https://doi.org/10.1016/0928-4257(94)90087-6 Get rights and content

Abstract

The inhibitory glycine receptor (GlyR) mediates postsynaptic inhibition in spinal cord, brain stem and other regions of the vertebrate central nervous system. Biochemical and molecular approaches have identified different developmentally and regionally regulated GlyR isoforms that result from the differential expression of at least four genes coding for different variants of the ligand-binding α subunit. Molecular studies have allowed identification of GlyR subunit domains implicated in ligand binding, channel formation and receptor assembly. At the postsynaptic membrane, the GlyR colocalizes with a 93-kDa tubulin-binding peripheral membrane protein, gephyrin. Antisense inhibition of gephyrin expression prevents GlyR accumulation at postsynaptic membrane specialization. Thus gephyrin is essential for postsynaptic receptor topology.

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Synaptic changes underlying the strengthening of GABA/glycinergic connections in the developing lateral superior olive
2010, Neuroscience
Citation Excerpt :
In the anteroventral cochlear nucleus larger glycine receptor cluster size correlate with larger mIPSC amplitudes (Lim et al., 1999). In the spinal cord and other brain areas, a developmental switch of glycine receptor subunit has been observed, where initially α2 subunit is predominant, but later replaced by α1 subunit (Betz et al., 1994). At least in the rat LSO, this switch does not seem to occur because the level of mRNA for α2 subunit is very low at birth and remains low, while α1 mRNA level increases during the first two postnatal weeks (Piechotta et al., 2001).
Before hearing onset, the topographic organization of the auditory GABA/glycinergic pathway from the medial nucleus of the trapezoid body (MNTB) to the lateral superior olive (LSO) is refined by synaptic silencing and strengthening. The synaptic mechanisms underlying the developmental strengthening of maintained MNTB-LSO connections are unknown. Here we address this question using whole-cell recordings from LSO neurons in slices prepared from prehearing mice. Minimal and maximal stimulation techniques demonstrated that during the first two postnatal weeks, individual LSO neurons lose about 55% of their initial presynaptic MNTB partners while maintained single-fiber connections become about 14-fold stronger. Analysis of MNTB-evoked miniature events indicates that this strengthening is accompanied by a 2-fold increase in quantal amplitude. Strengthening is not caused by an increase in the probability of release because paired pulse ratios (PPRs) increased from 0.7 in newborn animals to 0.9 around hearing onset, indicating a developmental decrease rather than increase in release probability. In addition, a possible soma-dendritic relocation of MNTB input seems unlikely to underlie their strengthening as indicated by analysis of the rise times of synaptic currents. Taken together, we conclude that the developmental strengthening of MNTB-LSO connections is achieved by a 2-fold increase in quantal size and an 8-fold increase in quantal content.
Plasticity at glycinergic synapses in dorsal cochlear nucleus of rats with behavioral evidence of tinnitus
2009, Neuroscience
Fifteen percent to 35% of the United States population experiences tinnitus, a subjective “ringing in the ears”. Up to 10% of those afflicted report severe and disabling symptoms. Tinnitus was induced in rats using unilateral, 1 h, 17 kHz-centered octave-band noise (116 dB SPL) and assessed using a gap-startle method. The dorsal cochlear nucleus (DCN) is thought to undergo plastic changes suggestive of altered inhibitory function during tinnitus development. Exposed rats showed near pre-exposure auditory brainstem response (ABR) thresholds for clicks and all tested frequencies 16 weeks post-exposure. Sound-exposed rats showed significantly worse gap detection at 24 and 32 kHz 16 weeks following sound exposure, suggesting the development of chronic, high frequency tinnitus. Message and protein levels of α_1–3, and β glycine receptor subunits (GlyRs), and the anchoring protein, gephyrin, were measured in DCN fusiform cells 4 months following sound exposure. Rats with evidence of tinnitus showed significant GlyR α₁ protein decreases in the middle and high frequency regions of the DCN while α₁ message levels were paradoxically increased. Gephyrin levels showed significant tinnitus-related increases in sound-exposed rats suggesting intracellular receptor trafficking changes following sound exposure. Consistent with decreased α₁ subunit protein levels, strychnine binding studies showed significant tinnitus-related decreases in the number of GlyR binding sites, supporting tinnitus-related changes in the number and/or composition of GlyRs. Collectively, these findings suggest the development of tinnitus is likely associated with functional GlyR changes in DCN fusiform cells consistent with previously described behavioral and neurophysiologic changes. Tinnitus related GlyR changes could provide a unique receptor target for tinnitus pharmacotherapy or blockade of tinnitus initiation.
Age-related changes in glycine receptor subunit composition and binding in dorsal cochlear nucleus
2009, Neuroscience
Citation Excerpt :
The significant age-related decreases of GlyR α1 and α2 protein staining of fusiform cells in middle and high frequency areas of DCN may correlate directly with the loss of strychnine binding sites (Fig. 6), consistent with previous rat and mouse aging studies (Frostholm and Rotter, 1985, 1986; Glendenning and Baker, 1988; Milbrandt and Caspary, 1995). Strychnine receptor binding is thought to be α1 sensitive and binds to the interface between α and β subunits (Young and Snyder, 1973, 1974; Betz et al., 1994; Schmieden and Betz, 1995; Legendre, 2001; Grudzinska et al., 2005). Thus, strychnine binding depends on the presence of these two subunits (Milbrandt and Caspary, 1995; Krenning et al., 1998).
Age-related hearing loss, presbycusis, can be thought of, in part, as a slow progressive peripheral deafferentation. Previous studies suggest that certain deficits seen in presbycusis may partially result from functional loss of the inhibitory neurotransmitter glycine in dorsal cochlear nucleus (DCN). The present study assessed age-related behavioral gap detection changes and neurochemical changes of postsynaptic glycine receptor (GlyRs) subunits and their anchoring protein gephyrin in fusiform cells of young (7–11 months) and aged (28–33 months) Fischer brown Norway (FBN) rats. Aged rats showed significantly (20–30 dB) elevated auditory brainstem-evoked response thresholds across all tested frequencies and worse gap detection ability compared to young FBN rats. In situ hybridization and quantitative immunocytochemistry were used to measure GlyR subunit message and protein levels. There were significant age-related increases in the α₁ subunit message with significant age-related decreases in α₁ subunit protein. Gephyrin message and protein showed significant increases in aged DCN fusiform cells. The pharmacologic consequences of these age-related subunit changes were assessed using [³H] strychnine binding. In support of the age-related decrease of α₁ subunit protein levels in DCN, there was a significant age-related decrease in the total number of GlyR binding sites with no significant change in affinity. These age-related changes may reflect an effort to reestablish a homeostatic balance between excitation and inhibition impacting on DCN fusiform cells by downregulation of inhibitory function in the face of an age-related loss of peripheral input. Age-related decrease in presynaptic glycine release results in altered subunit composition and this may correlate with loss of temporal coding of the aged fusiform cell in DCN. The previously reported role for gephyrin in retrograde intracellular receptor subunit trafficking could contribute to the α₁ decrease in the face of increased message.
Genistein directly blocks glycine receptors of rat neurons freshly isolated from the ventral tegmental area
2003, Neuropharmacology
The effects of tyrosine kinase inhibitors on the glycine-induced current (I_Gly) were studied in rat neurons freshly isolated from the ventral tegmental area (VTA). Genistein reversibly and concentration-dependently depressed I_Gly, with an IC₅₀ of 13 μM. Preincubation with genistein had no effect on I_Gly, indicating that genistein is effective only when glycine is bound to the receptor and channels are most likely open. Genistein depressed maximum I_Gly without significantly changing the EC₅₀ for glycine. Genistein-induced inhibition of I_Gly was sensitive to membrane voltage, being greater at positive membrane potentials. A kinetic analysis indicated that genistein lengthens the time constant of I_Gly activation, but has no effect on deactivation or desensitization. When genistein was rapidly washed out, a transient rebound current probably reflected a faster dissociation of genistein, with respect to glycine. Results of competition experiments suggest that genistein acts on the same region of the glycine receptor as picrotoxin. Daidzein, an analog of genistein that does not act on protein kinases, also inhibited I_Gly. Co-application of lavendustin A, a specific inhibitor of tyrosine kinase, had no effect on I_Gly. Our results extend to neurons isolated from the VTA, the previous finding that genistein directly inhibits glycine receptors of hypothalamic brain slices.
Astrocytic receptors and second messenger systems
2003, Advances in Molecular and Cell Biology
Citation Excerpt :
Functional glycine receptors have been detected on astrocytes. These receptors mediate a Cl− conductance (Betz et al., 1994) and show similarities with the GABAA receptor. The monoamines noradrenaline and dopamine share a common biosynthetic pathway that starts with the amino acid tyrosine.
This chapter reviews that glial cells support neuronal activity and create opportunities for the dynamics and plasticity of the central nervous system (CNS). After injury, metabolic disturbance or toxic influence, and in degenerative disease, glial cell reactions protect neurons and facilitate rebuilding. It explores that astrocytes is the most numerous glial cells in the CNS and express a large number of receptors for neurotransmitters, peptides, purines, cytokines, and other neuroactive substances; many of them previously thought to be present only on neurons. These astrocytic receptors are coupled to G-proteins or ion channels, to intracellular protein kinases, or associated with mitochondria or cell nuclei. Even if most evidence for the existence of this extensive repertoire of functional astrocytic receptors comes from in vitro studies, convincing studies have demonstrated the presence of astrocytic receptor groups also in the intact nervous system. The chapter also discusses that the astrocytic receptors and the signaling systems, with which they are associated, set the stage for extensive neuronal–glial signaling that in turn is essential for neuroplasticity both in the intact nervous system, and after trauma and in degenerative disease. Pharmacological manipulation of astrocytic receptor systems might provide a new strategy to reinforce the reparative processes within the CNS after injury or in disease.
Why glycine transporters have different stoichiometries
2002, FEBS Letters
In the brain, neurons and glial cells compete for the uptake of the fast neurotransmitters, glutamate, GABA and glycine, through specific transporters. The relative contributions of glia and neurons to the neurotransmitter uptake depend on the kinetic properties, thermodynamic coupling and density of transporters but also on the intracellular metabolization or sequestration of the neurotransmitter. In the case of glycine, which is both an inhibitory transmitter and a neuromodulator of the excitatory glutamatergic transmission as a co-agonist of N-methyl D-aspartate receptors, the glial (GlyT1b) and neuronal (GlyT2a) transporters differ at least in three aspects: (i) stoichiometries, (ii) reverse uptake capabilities and (iii) pre-steady-state kinetics. A 3 Na⁺/1 Cl⁻/gly stoichiometry was established for GlyT2a on the basis of a 2 charges/glycine flux ratio and changes in the reversal potential of the transporter current as a function of the extracellular glycine, Na⁺ and Cl⁻ concentrations. Therefore, the driving force available for glycine uphill transport in neurons is about two orders of magnitude larger than for glial cells. In addition, GlyT2a shows a severe limitation for reverse uptake, which suggests an essential role of GlyT2a in maintaining a high intracellular glycine pool, thus facilitating the refilling of synaptic vesicles by the low affinity, low specificity vesicular transporter VGAT/VIAAT. In contrast, the 2 Na⁺/1 Cl⁻/gly stoichiometry and bi-directional transport properties of GlyT1b are appropriate for the control of the extracellular glycine concentration in a submicromolar range that can modulate N-methyl D-aspartate receptors effectively. Finally, analysis of the pre-steady-state kinetics of GlyT1b and GlyT2a revealed that at the resting potential neuronal transporters are preferentially oriented outward, ready to bind glycine, which suggests a kinetic advantage in the uptake contest.

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Structure, diversity and synaptic localization of inhibitory glycine receptors

Abstract

Brain Res

Neuron

Neuron

Trends Neurosci

Biochem Biophys Res Commun

Neuron

Neuron

J Biol. Chem

Brain res

Neuron

J Biol Chem

FEBS Lett

Neuron

Biochim Biophys Acta

J Biol Chem

J Biol Chem

Neuron

Neuron

Cell 72/Neuron

Heterogeneity of glycine receptors and their RNAs in rat brain and spinal cord

Science

Cloning of a glycine receprot sybtype expressed in rat brain and spinal cord during a specific period of neuronal development

FEBS Lett

The glycine receptor deficiency of the mutant mouse spastic: evidence for normal glycine receptor structure and localization

J Neurosci

Glycine receptor heterogeneity in rat spinal cord during postnatal development

EMBO J

Residues within transmembrane segment M2 determine chloride conductance of glycine receptor homo- and hetero-oligomers

EMBO J