Glutamate Receptors In Neurons Research Articles

Neuronal Glutamate and GABAA Receptor Function in Health and Disease

Glutamate and GABA (gamma-aminobutyric acid) are the predominant excitatory and inhibitory neurotransmitters in the mammalian CNS (central nervous system) respectively, and as such have undergone intense investigation. Given their predominance, it is no wonder that the reciprocal receptors for these neurotransmitters have attracted so much attention as potential targets for the promotion of health and the treatment of disease. Indeed, dysfunction of these receptors underlies a number of well-characterized neuropathological conditions such as anxiety, epilepsy and neurodegenerative diseases. Although intrinsically linked, the glutamatergic and GABAergic systems have, by and large, been investigated independently, with researchers falling into the 'excitatory' or 'inhibitory' camps. Around 70 delegates gathered at the University of St Andrews for this Biochemical Society Focused Meeting aimed at bringing excitation and inhibition together. With sessions on behaviour, receptor structure and function, receptor trafficking, activity-dependent changes in gene expression and excitation/inhibition in disease, the meeting was the ideal occasion for delegates from both backgrounds to interact. This issue of Biochemical Society Transactions contains papers written by those who gave oral presentations at the meeting. In this brief introductory review, I put into context and give a brief overview of these contributions.

Mechanisms of Modulation of AMPA-Induced Na+-Activated K+ Current by mGluR1

Na(+)-activated K(+) (K(Na)) channels can be activated by Na(+) influx via ionotropic receptors and play a role in shaping synaptic transmission. In expression systems, K(Na) channels are modulated by G protein-coupled receptors, but such a modulation has not been shown for the native channels. In this study, we examined whether K(Na) channels coupled to AMPA receptors are modulated by metabotropic glutamate receptors (mGluRs) in lamprey spinal cord neurons. Activation of mGluR1 strongly inhibited the AMPA-induced K(Na) current. However, when intracellular Ca(2+) was chelated with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), the K(Na) current was enhanced by mGluR1. Activation of protein kinase C (PKC) mimicked the inhibitory effect of mGluR1 on the K(Na) current. Blockade of PKC prevented the mGluR1-induced inhibition of the K(Na) current, but did not affect the enhancement of the current seen in BAPTA. Together these results suggest that mGluR1 can differentially modulate AMPA-induced K(Na) current in a Ca(2+)- and PKC-dependent manner.

Role of Neuronal NMDA and non-NMDA Glutamate Receptors in Medial Vestibular Nucleus in the Regulation of Respiratory Rhythmogenesis in Newborn Rats In Vitro

We studied isolated pontobulbospinal preparations from newborn rat brain. In the early postnatal period, the rostral part of the medial vestibular nucleus produces a potent inhibitory effect on neuronal structures of the bulbar respiratory center via the glutamatergic system. Microinjection of L-glutamate (50 mmol/liter) into the rostral part of the vestibular nucleus completely blocks respiratory rhythmogenesis in 0-1-day-old rat pups and reduced the frequency of generation of inspiratory discharges in 2-3-day-old rats from 8.42+/-0.68 to 2.68+/-0.32 min(-1). It was found that the leading role in the mechanism of glutamatergic modulation of the respiratory rhythmogenesis by neurons of the medial vestibular nucleus is played by NMDA and, to a lesser extent, non-NMDA glutamate receptors.

Long-term exposure to endogenous levels of tributyltin decreases GluR2 expression and increases neuronal vulnerability to glutamate

Tributyltin (TBT), an endocrine-disrupting chemical, has been used commercially as a heat stabilizer, agricultural pesticide and component of antifouling paints. In this study, we investigated the effect of long-term exposure to endogenous levels of TBT on neuronal glutamate receptors. Cultured rat cortical neurons were exposed to 1–50 nM TBT for 9 days (from day 2 to day 10 in vitro). The number of neurons was reduced by long-term exposure to 50 nM TBT, but not to 1–20 nM TBT. Long-term exposure to 20 nM TBT decreased the mRNA expression of glutamate receptors NR1, NR2A, GluR1 and GluR2, and increased that of NR2B, GluR3 and GluR4. GluR2 protein was also reduced by long-term exposure to TBT. Because AMPA receptor lacking GluR2 exhibits Ca 2+ permeability, we investigated whether Ca 2+ influx or glutamate toxicity was affected. Indeed, glutamate-induced Ca 2+ influx was increased in TBT-treated neurons. Consistent with this, neurons became more susceptible to glutamate toxicity as a result of long-term exposure to TBT and this susceptibility was abolished by an antagonist of GluR2-lacking AMPA receptor. Thus, it is suggested that long-term exposure to endogenous levels of TBT induces a decrease of GluR2 protein, causing neurons become more susceptible to glutamate toxicity.

Excitotoxic Death of Retinal NeuronsIn VivoOccurs via a Non-Cell-Autonomous Mechanism

The central hypothesis of excitotoxicity is that excessive stimulation of neuronal NMDA-sensitive glutamate receptors is harmful to neurons and contributes to a variety of neurological disorders. Glial cells have been proposed to participate in excitotoxic neuronal loss, but their precise role is defined poorly. In this in vivo study, we show that NMDA induces profound nuclear factor kappaB (NF-kappaB) activation in Müller glia but not in retinal neurons. Intriguingly, NMDA-induced death of retinal neurons is effectively blocked by inhibitors of NF-kappaB activity. We demonstrate that tumor necrosis factor alpha (TNFalpha) protein produced in Müller glial cells via an NMDA-induced NF-kappaB-dependent pathway plays a crucial role in excitotoxic loss of retinal neurons. This cell loss occurs mainly through a TNFalpha-dependent increase in Ca(2+)-permeable AMPA receptors on susceptible neurons. Thus, our data reveal a novel non-cell-autonomous mechanism by which glial cells can profoundly exacerbate neuronal death following excitotoxic injury.

Nitrous oxide and the inhibitory synaptic transmission in rat dorsal horn neurons

The analgesic effect of nitrous oxide (N 2O) is thought to depend on noradrenaline release in the spinal cord following activation of descending inhibitory neurons. In addition to this indirect facilitation of inhibition in the spinal cord, we previously showed direct inhibition of glutamate receptors in dorsal horn neurons by N 2O. Since general anesthetics could possibly affect excitatory and/or inhibitory components of synaptic transmission, we sought to evaluate the direct effect of N 2O on inhibitory transmission in spinal cord neurons. Using whole-cell patch-clamp recording from rat transversal spinal cord slices, we investigated the actions of 50% N 2O and 0.5% isoflurane (both 0.3 rat MAC; minimum alveolar concentration) on exogenously applied γ-aminobutyric acid (GABA)- and glycine-induced currents in rat dorsal horn lamina II neurons. The amplitudes and integrated areas of GABA- and glycine-induced currents were not significantly affected by N 2O, but were increased in the presence of isoflurane. N 2O did not affect the amplitude, frequency or decay time probability distribution of either GABA or glycine receptor-mediated miniature postsynaptic currents. We further sought to determine the effect of N 2O on focal stimulation-evoked synaptic currents mediated by GABA and glycine receptors, and found no effect in the majority of neurons. These and other findings suggest that N 2O has a discrete action in the spinal cord, distinct from the effects of the volatile anesthetics, consisting of inhibition of excitation in SG neurons through an action on ionotropic glutamatergic receptors and potentiation of inhibition through the descending noradrenergic system.

Development of glutamate receptors in auditory neurons from long‐term organotypic cultures of the embryonic chick hindbrain

We used long-range organotypic cultures of auditory nuclei in the chick hindbrain to test the development of glutamate receptor activity in auditory neurons growing in a tissue environment that includes early deprivation of peripheral glutamatergic input, subsequent to removal of the otocyst. Cultures started at embryonic day (E)5, and lasted from 6 h to 15 days. Neuronal migration, clustering and axonal extension from the nucleus magnocellularis (NM) to the nucleus laminaris (NL) partially resembled events in vivo. However, the distinctive laminar organization of the NL was not observed. Glutamate receptor (GluR) activity was tested with optical recordings of intracellular Ca2+ in the NM. alpha-Amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (AMPA)/kainate receptors had Ca2+ responses with a time course similar to that in control slices. Peak amplitude, however, was significantly lower. N-methyl-D-aspartate (NMDA)-mediated Ca2+ responses were higher in 2-day cultures (E5 + 2d) than in E7 explant controls, returning later to control values. Metabotropic GluRs did not elicit Ca2+ responses at standard agonist doses. Blocking NMDA or AMPA/kainate receptors with specific antagonists for 10 days in culture did not limit neuronal survival. Blocking metabotropic GluRs resulted in complete neuronal loss. Thus, ionotropic GluRs are not required for NM neuronal survival. However, their activity during development is affected when neurons grow in an in vitro environment that includes prevention of arrival of peripheral glutamatergic input.

Developmental regulation of ionotropic glutamate receptors in central vestibular neurons in rats

Developmental regulation of ionotropic glutamate receptors in central vestibular neurons in rats

Developmental maturation of ionotropic glutamate receptor subunits in rat vestibular nuclear neurons responsive to vertical linear acceleration

We investigated the maturation profile of subunits of ionotropic glutamate receptors in vestibular nuclear neurons that were activated by sinusoidal linear acceleration along the vertical plane. The otolithic origin of Fos expression in these neurons was confirmed as a marker of functional activation when labyrinthectomized and/or stationary control rats contrasted by showing sporadically scattered Fos-labeled neurons in the vestibular nuclei. By double immunohistochemistry for Fos and one of the receptor subunits, otolith-related neurons that expressed either alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate or N-methyl-d-aspartate subunits were first identified in the medial vestibular nucleus, spinal vestibular nucleus and Group x by postnatal day (P)7, and in the lateral vestibular nucleus and Group y by P9. No double-labeled neurons were found in the superior vestibular nucleus. Within each vestibular subnucleus, these double-labeled neurons constituted approximately 90% of the total Fos-labeled neurons. The percentage of Fos-labeled neurons expressing the GluR1 or NR2A subunit showed developmental invariance in all subnuclei. For Fos-labeled neurons expressing the NR1 subunit, similar invariance was observed except that, in Group y, these neurons decreased from P14 onwards. For Fos-labeled neurons expressing the GluR2, GluR2/3, GluR4 or NR2B subunit, a significant decrease was found by the adult stage. In particular, those expressing the GluR4 subunit showed a two- to threefold decrease in the medial vestibular nucleus, spinal vestibular nucleus and Group y. Also, those expressing the NR2B subunit showed a twofold decrease in Group y. Taken together, the postsynaptic expression of ionotropic glutamate receptor subunits in different vestibular subnuclei suggests that glutamatergic transmission within subregions plays differential developmental roles in the coding of gravity-related vertical spatial information.

Elevated glycogen synthase kinase-3 activity in Fragile X mice: Key metabolic regulator with evidence for treatment potential

Significant advances have been made in understanding the underlying defects of and developing potential treatments for Fragile X syndrome (FXS), the most common heritable mental retardation. It has been shown that neuronal metabotropic glutamate receptor 5 (mGluR5)-mediated signaling is affected in FX animal models, with consequent alterations in activity-dependent protein translation and synaptic spine functionality. We demonstrate here that a central metabolic regulatory enzyme, glycogen synthase kinase-3 (GSK3) is present in a form indicating elevated activity in several regions of the FX mouse brain. Furthermore, we show that selective GSK3 inhibitors, as well as lithium, are able to revert mutant phenotypes of the FX mouse. Lithium, in particular, remained effective with chronic administration, although its effects were reversible even when given from birth. The combination of an mGluR5 antagonist and GSK3 inhibitors was not additive. Instead, it was discovered that mGluR5 signaling and GSK3 activation in the FX mouse are coordinately elevated, with inhibition of mGluR5 leading to inhibition of GSK3. These findings raise the possibility that GSK3 is a fundamental and central component of FXS pathology, with a substantial treatment potential.

Neuron-glia crosstalk gets serious: role in pain hypersensitivity.

Recent studies show that peripheral injury activates both neuronal and nonneuronal or glial components of the peripheral and central cellular circuitry. The subsequent neuron-glia interactions contribute to pain hypersensitivity. This review will briefly discuss novel findings that have shed light on the cellular mechanisms of neuron-glia interactions in persistent pain. Two fundamental questions related to neuron-glia interactions in pain mechanisms have been addressed: what are the signals that lead to central glial activation after injury and how do glial cells affect central nervous system neuronal activity and promote hyperalgesia? Evidence indicates that central glial activation depends on nerve inputs from the site of injury and release of chemical mediators. Hematogenous immune cells may migrate to/infiltrate the brain and circulating inflammatory mediators may penetrate the blood-brain barrier to participate in central glial responses to injury. Inflammatory cytokines such as interleukin-1beta released from glia may facilitate pain transmission through its coupling to neuronal glutamate receptors. This bidirectional neuron-glia signaling plays a key role in glial activation, cytokine production and the initiation and maintenance of hyperalgesia. Recognition of the contribution of the mutual neuron-glia interactions to central sensitization and hyperalgesia prompts new treatment for chronic pain.

Glutamate neurotoxicity: Perturbations of ionic homeostasis, mitochondrial dysfunction, and changes in cell functioning

The hyperstimulation of neuronal glutamate receptors during different pathologies results in CNS injuries. This review summarizes the data concerning changes in the concentrations of sodium, potassium, calcium, and protons in the cytosol and mitochondrial matrix of cultured neurons subjected to prolonged glutamate treatment. Development of delayed calcium deregulation (DCD) plays a leading role in the initiation of neuronal death during toxic glutamate treatment. We review the mechanisms of DCD development, the role of mitochondria in the regulation of neuronal calcium homeostasis during stimulation of glutamate receptors, and intracellular events that accompany DCD and may result in neuronal death.

Chemokine CXCL8 modulates GluR1 phosphorylation

The chemokine interleukin 8/CXCL8 induces the phosphorylation of the GluR1 subunit of the AMPA-type glutamate receptor in neurons and transfected HEK cells, on both serine 845 (S845) and 831 (S831) residues. We previously described that CXCL8 receptor CXCR2 and GluR1 co-precipitate and that GluR1/CXCR2 co-expression both in HEK cells and neurons impairs CXCL8-induced cell migration. Here we show that replacement of S845 with Ala (A), but not with Glu (E), strongly reduces GluR1/CXCR2 interaction and abolishes the impairment of CXCL8-induced cell migration. Considered together our findings point to the phosphorylated state of S845GluR1 as a determinant of GluR1–CXCR2 physical coupling.

Coupling between neuronal nitric oxide synthase and glutamate receptor 6–mediated c-Jun N-terminal kinase signaling pathway via S-nitrosylation contributes to ischemia neuronal death

S-nitrosylation, as a post-translational protein modification, recently has been paid more and more attention in stroke research. S-nitrosylation regulates protein function by the mechanisms of covalent attachment that control the addition or the removal of nitric oxide (NO) from a cysteine thiol. The derivation of NO is established by the demonstration that, in cerebral neurons, NO mainly generates from neuronal nitric oxide synthase (nNOS) during the early stages of reperfusion. In the past researches, we demonstrate that global ischemia–reperfusion facilitates the activation of glutamate receptor 6 (GluR6) –mediated c-Jun N-terminal kinase (JNK) signaling pathway. The objective of this study is primarily to determine, during the early stages of reperfusion in rat four-vessel occlusion (4-VO) ischemic model, whether nNOS-derived NO affects the GluR6-mediated JNK signaling route via S-nitrosylation which is performed mainly by the biotin switch assay. Here, we show that administration of 7-nitroindazole, an inhibitor of nNOS, or ketamine, an antagonist of N-methyl- d-aspartate receptor (NMDAR), diminishes the increased S-nitrosylation of GluR6 induced by cerebral ischemia–reperfusion. In contrast, 2-amion-5,6-dihydro-6-methyl-4H-1,3-thiazine, an inhibitor of inducible NO synthase does not affect S-nitrosylation of GluR6. Moreover, treatment with sodium nitroprusside (SNP), an exogenous NO donor, increases the S-nitrosylation and phosphorylation of nNOS, leading to the attenuation of the increased S-nitrosylation of GluR6 and the assembling of GluR6• postsynaptic density protein 95 (PSD95)• mixed lineage kinase 3 (MLK3) signaling module induced by cerebral ischemia–reperfusion. The results also show that GluR6 downstream MLK3• mitogen activated protein kinase kinase 4/7• JNK signaling module and nuclear or non-nuclear apoptosis pathways are involved in the above signaling route. However, dithiothreitol (DTT) antagonizes the neuroprotection of SNP. Treatment with DTT alone, as a negative control, prevents S-nitrosylation of proteins, which indicates the existence of endogenously produced S-nitrosylation. These data suggest that GluR6 is S-nitrosylated by endogenous NO in cerebral ischemia–reperfusion, which is possibly correlated with NMDAR• PSD95• nNOS signaling module, and further activates GluR6• PSD95• MLK3 signaling module and JNK signaling pathway. In contrast, exogenous NO donor antagonizes the above action of endogenous NO generated from nNOS. Thus, our results provide the coupling of nNOS with GluR6 by S-nitrosylation during the early stages of ischemia–reperfusion, which can be a new approach for stroke therapy.

Morphometric analysis of the AMPA-type neurons in the Deiters vestibular complex of the chick brain

Chicken ( Gallus gallus) brains were used to investigate the typology and the immunolabel pattern for the subunits composing the AMPA-type glutamate receptors (GluR) of hindbrain neurons of the dorsal (dND) and ventral nuclei (vND) of the Deiter's vestibular complex (CD), which is the avian correspondent of the lateral vestibular nucleus (LVN) of mammals. Our results revealed that neurons of both divisions were poor in GluR1. The vND, the GluR2/3+ and GluR4+ label presented no area or neuronal size preference, although most neurons were around 75%. The dND neurons expressing GluR2/3 are primarily around 85%, medium to large-sized 85%, and predominantly 60% located in the medial portion of the rostral pole and in the lateral portion of the caudal pole. The majority of dND neurons containing GluR4 are also around 75%, larger (70% are large and giant), exhibiting a distribution that seems to be complementary to that of GluR2/3+ neurons. This distinct arrangement indicates functional differences into and between the DC nuclei, also signaling that such variation could be attributed to the diverse nature of the subunit composition of the GluRs. Discussion addresses the morphological and functional correlation of the avian DC with the LVN of mammals in addition to the high morphological correspondence, To include these data into the modern comparative approach we propose to adopt a similar nomenclature for the avian divisions dND and vND that could be referred as dLVN and vLVN.

Long-lasting dephosphorylation of connexin 43 in acute seizures is regulated by NMDA receptors in the rat cerebral cortex

Gap junctions consisting of connexin 43 subunits provide intercellular communication between astrocytes, contributing to their function of maintaining the central nervous system (CNS) microenvironment. Magnetic resonance imaging (MRI) studies demonstrate prolonged astrocyte swelling related to seizures, while in vitro studies show the disruption of intercellular coupling and the cytotoxic swelling of astrocytes in seizure-equivalent environments. We examined the relation between astrocyte swelling and connexin 43 regulation using an in vivo seizure model. Generalised tonic-clonic convulsions were induced in adult rats using intraperitoneally (i.p.)-administered 4-aminopyridine (4-AP). The expression of connexin 43 mRNA and protein at 1, 3 and 24 h after seizure induction was measured. Astrocytic swelling was assessed at the same time points using transmission electron microscopy. mRNA and protein levels remained unaltered at all time periods. However, a significant (≈50%) reduction was found in the amount of phosphorylated (P1, P2) to unphosphorylated (NP) forms of connexin 43 at 3 h. The amount increased thereafter, but was still significantly lower than in the controls at 24 h post-seizure. Simultaneously, marked astrocytic swelling was measured in the neocortex. Pre-treatment with N-methyl-D-aspartate (NMDA) receptor antagonist dizocilpine maleate (MK-801) resulted in the amelioration of seizure symptoms and the prevention of connexin 43 dephosphorylation, as well as significantly reduced astrocytic swelling. Dephosphorylation of connexin 43 was shown to reduce astrocytic gap junction (GJ) permeability. Our results therefore suggest that, during acute seizures, a prolonged inhibition of intercellular coupling develops in the astrocyte network. This accounts for the long-lasting astrocyte swelling observed, and potentially impairs buffering function. The results also imply that this uncoupling is regulated through neuronal and/or glial NMDA-type glutamate receptors.

Chemical Two-Photon Uncaging

INTRODUCTIONChemical two-photon uncaging is useful for a wide range of applications, including both mapping of receptor location and localized photostimulation of neurons via activation of excitatory glutamate receptors. Experimental preparations could include brain slices, cultured neurons, and, among other possibilities, whole brains in vivo. This protocol documents the utility of chemical two-photon uncaging in examining glutamate receptors of pyramidal neurons in hippocampal slices.

Sequential Postsynaptic Maturation Governs the Temporal Order of GABAergic and Glutamatergic Synaptogenesis in Rat Embryonic Cultures

Sequential formation of GABAergic and glutamatergic synapses is thought to be crucial for constructing the stereotypic neural networks during brain development. However, why GABAergic synapses are formed earlier than glutamatergic synapses in the developing brain is not well understood. We used electrophysiology and fluorescence imaging to study GABAergic and glutamatergic synaptogenesis in embryonic hypothalamic cultures, which contain approximately 40% GABAergic and approximately 60% glutamatergic neurons. The newly dissociated embryonic hypothalamic neurons contained a significant pool of functional GABA(A) receptors but a very low level of glutamate receptors. Within the first week of culture, the time course of GABAergic synaptogenesis in embryonic neurons coincided with that of presynaptic vesicle cycling, but both measurements lagged behind the detection of functional GABA(A) receptors. Remarkably, the GABA(A) receptors of newly dissociated embryonic neurons can be rapidly clustered into postsynaptic apparatus and generate functional synaptic currents within 4-6 h when cocultured with mature neurons. Consistent with earlier expression of GABA(A) receptors in immature neurons, synaptic GABAergic events were always detected before the onset of glutamatergic events in both purely embryonic and heterochronic cultures. Interestingly, overexpression of glutamate receptors in embryonic neurons not only increased whole-cell glutamate currents but also significantly increased the frequency of excitatory synaptic events. We conclude that the sequential formation of GABAergic and glutamatergic synapses in immature neurons is likely governed by a sequential expression of GABA(A) and glutamate receptors during neuronal development.

Amyloid β Protein Modulates Glutamate-Mediated Neurotransmission in the Rat Basal Forebrain: Involvement of Presynaptic Neuronal Nicotinic Acetylcholine and Metabotropic Glutamate Receptors

Amyloid beta (Abeta) protein, a 39-43 amino acid peptide deposited in brains of individuals with Alzheimer's disease (AD), has been shown to interact directly with a number of receptor targets including neuronal nicotinic acetylcholine receptors (nAChRs) and glutamate receptors. In this study, we investigated the synaptic effects of Abeta(1-42) on glutamate-mediated neurotransmission in the diagonal band of Broca (DBB), a cholinergic basal forebrain nucleus. Glutamatergic miniature EPSCs (mEPSCs) were recorded using whole-cell patch-clamp recordings from identified cholinergic DBB neurons in rat forebrain slices. In 54% of DBB neurons, bath application of Abeta(1-42) (100 nM), but not Abeta(42-1) (inverse fragment), significantly increased the frequency of mEPSCs without affecting amplitude or kinetic parameters (rise or decay time). In 32% of DBB neurons, bath application of Abeta(1-42) significantly decreased only the frequency but not amplitude of mEPSCs. Application of dihydro-beta-erythroidine (DHbetaE) (an antagonist for the alpha4beta2 subtype of nAChRs) but not alpha-bungarotoxin (an antagonist for the alpha7 subtype of nAChRs) blocked Abeta(1-42)-mediated increases in mEPSC frequency. The Abeta(1-42)-mediated increase in glutamatergic transmission is thus presynaptic and mediated via non-alpha7 AChRs. In contrast, Abeta(1-42)-mediated decreases in mEPSC frequency could not be antagonized by either DHbetaE or alpha-bungarotoxin. However, the Abeta(1-42)-evoked depression in mEPSC frequency was antagonized by (RS)-alpha-methyl-4-carboxyphenyglycine, a nonselective group I/II metabotropic glutamate receptor antagonist. These observations provide further insight into the mechanisms whereby Abeta affects synaptic function in the brain and may be relevant in the context of synaptic failure observed in AD.

Co-expression of myosin II regulatory light chain and the NMDAR1 subunit in neonatal and adult mouse brain

Movement of glutamate receptors in neurons likely involves direct and indirect association of receptor subunits with microtubule- and actin-based motor proteins. We have previously shown that myosin II regulatory light chain (RLC) binds directly to subunits of the NMDA-type glutamate receptor (NR), suggesting that NMDA receptors are closely associated with a myosin II motor complex. Using a polyclonal antibody predicted to recognize all RLC isoforms previously described in rodent brain, we report the expression of RLC and the NR1 subunit in cortex, hippocampus and cerebellum of postnatal day 0 (P0) and adult mouse. Although myosin RLC was not exclusively localized with NR1 by immunohistochemistry, co-staining was striking in the neuronal soma of deep cortical neurons and Purkinje neurons of the cerebellum which showed a punctate, perinuclear pattern of immunoreactivity. These neuronal populations were identified using a monoclonal antibody directed against a nuclear-specific, transcriptional repressor, chicken ovalbumin upstream promoter-transcription factor (COUP-TF)-interacting protein 2 (CTIP2). Co-expression of NR1 and a myosin II motor was validated using an isoform specific anti-nonmuscle myosin II-B heavy chain (NMHC II-B) antibody. Our findings support the idea that there is regional heterogeneity in the molecular composition of the NMDA receptor-associated cytoskeleton, and suggest that NR subunits may be associated with an actin-based, myosin II-B motor within the endomembrane system of some neuronal populations. Differential staining patterns observed with light and heavy chain antibodies, however, suggest that there is also heterogeneity in the composition of myosin II complexes in brain.