Kainate Receptor Agonist Research Articles

Disposable facemask waste combustion emits neuroactive smoke particulate matter

Tremendous deposits of disposable medical facemask waste after the COVID-19 pandemic require improvement of waste management practice according to WHO report 2022, moreover facemasks are still in use around the world to protect against numerous airborne infections. Here, water-suspended smoke preparations from the combustion of disposable medical facemasks (polypropylene fibers) were collected; size, zeta potential, surface groups of smoke particulate matter were determined by dynamic light scattering, FTIR and Raman spectroscopy, and their optical properties were characterized. Neurochemical study using nerve terminals isolated from rat cortex revealed a significant decrease in the initial rate of the uptake/accumulation of excitatory and inhibitory neurotransmitters, L-[14C]glutamate and [3H]GABA, and exocytotic release, and also an increase in the extracellular level of these neurotransmitters. Fluorescent measurements revealed that ROS generation induced by hydrogen peroxide and glutamate receptor agonist kainate decreased in nerve terminals. A decrease in the membrane potential of nerve terminals and isolated neurons, the mitochondrial potential and synaptic vesicle acidification was also shown. Therefore, accidental or intentional utilization of disposable medical facemask waste by combustion results in the release of neuroactive ultrafine particulate matter to the environment, thereby contributing to plastic-associated pollution of air and water resources and neuropathology development and expansion.

An In Vivo Electroencephalographic Analysis of the Effect of Riluzole against Limbic and Absence Seizure and Comparison with Glutamate Antagonists.

Riluzole (RLZ) has demonstrated neuroprotective effects in several neurological disorders. These neuroprotective effects seem to be mainly due to its ability to inhibit the excitatory glutamatergic neurotransmission, acting on different targets located both at the presynaptic and postsynaptic levels. In the present study, we evaluated the effects of Riluzole (RLZ) against limbic seizures, induced by AMPA, kainate, and NMDA receptor agonists in Sprague-Dawley rats, and in a well-validated genetic model of absence epilepsy, the WAG/Rij rat. Furthermore, in this latter model, we also studied the effect of RLZ in co-administration with the competitive NMDA receptor antagonist, CPP, or the non-competitive AMPA receptor antagonist, THIQ-10c, on spike-wave discharges (SWDs) in WAG/Rij rats, to understand the potential involvement of AMPA and NMDA receptors in the anti-absence effect of RLZ. In Sprague-Dawley rats, RLZ pretreatment significantly reduced the limbic seizure severity induced by glutamatergic agonists, suggesting an antagonism of RLZ mainly on NMDA rather than non-NMDA receptors. RLZ also reduced SWD parameters in WAG/Rij rats. Interestingly, the co-administration of RLZ with CPP did not increase the anti-absence activity of RLZ in this model, advocating a competitive effect on the NMDA receptor. In contrast, the co-administration of RLZ with THIQ-10c induced an additive effect against absence seizure in WAG/Rij rats. these results suggest that the antiepileptic effects of RLZ, in both seizure models, can be mainly due to the antagonism of the NMDA glutamatergic receptors.

An Ene-reaction Ene-ables an α-Kainic Acid Synthesis

Key words ene reaction - kainate receptor agonists - Jones’ reagent

Willardiine and Its Synthetic Analogues: Biological Aspects and Implications in Peptide Chemistry of This Nucleobase Amino Acid

Willardiine is a nonprotein amino acid containing uracil, and thus classified as nucleobase amino acid or nucleoamino acid, that together with isowillardiine forms the family of uracilylalanines isolated more than six decades ago in higher plants. Willardiine acts as a partial agonist of ionotropic glutamate receptors and more in particular it agonizes the non-N-methyl-D-aspartate (non-NMDA) receptors of L-glutamate: ie. the α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) and kainate receptors. Several analogues and derivatives of willardiine have been synthesised in the laboratory in the last decades and these compounds show different binding affinities for the non-NMDA receptors. More in detail, the willardiine analogues have been employed not only in the investigation of the structure of AMPA and kainate receptors, but also to evaluate the effects of receptor activation in the various brain regions. Remarkably, there are a number of neurological diseases determined by alterations in glutamate signaling, and thus, ligands for AMPA and kainate receptors deserve attention as potential neurodrugs. In fact, similar to willardiine its analogues often act as agonists of AMPA and kainate receptors. A particular importance should be recognized to willardiine and its thymine-based analogue AlaT also in the peptide chemistry field. In fact, besides the naturally-occurring short nucleopeptides isolated from plant sources, there are different examples in which this class of nucleoamino acids was investigated for nucleopeptide development. The applications are various ranging from the realization of nucleopeptide/DNA chimeras for diagnostic applications, and nucleoamino acid derivatization of proteins for facilitating protein-nucleic acid interaction, to nucleopeptide-nucleopeptide molecular recognition for nanotechnological applications. All the above aspects on both chemistry and biotechnological applications of willardine/willardine-analogues and nucleopeptide will be reviewed in this work.

Reelin Is Required for Maintenance of Granule Cell Lamination in the Healthy and Epileptic Hippocampus.

One characteristic feature of mesial temporal lobe epilepsy is granule cell dispersion (GCD), a pathological widening of the granule cell layer in the dentate gyrus. The loss of the extracellular matrix protein Reelin, an important positional cue for neurons, correlates with GCD formation in MTLE patients and in rodent epilepsy models. Here, we used organotypic hippocampal slice cultures (OHSC) from transgenic mice expressing enhanced green fluorescent protein (eGFP) in differentiated granule cells (GCs) to monitor GCD formation dynamically by live cell video microscopy and to investigate the role of Reelin in this process. We present evidence that following treatment with the glutamate receptor agonist kainate (KA), eGFP-positive GCs migrated mainly toward the hilar region. In the hilus, Reelin-producing neurons were rapidly lost following KA treatment as shown in a detailed time series. Addition of recombinant Reelin fragments to the medium effectively prevented the KA-triggered movement of eGFP-positive GCs. Placement of Reelin-coated beads into the hilus of KA-treated cultures stopped the migration of GCs in a distance-dependent manner. In addition, quantitative Western blot analysis revealed that KA treatment affects the Reelin signal transduction pathway by increasing intracellular adaptor protein Disabled-1 synthesis and reducing the phosphorylation of cofilin, a downstream target of the Reelin pathway. Both events were normalized by addition of recombinant Reelin fragments. Finally, following neutralization of Reelin in healthy OHSC by incubation with the function-blocking CR-50 Reelin antibody, GCs started to migrate without any direction preference. Together, our findings demonstrate that normotopic position of Reelin is essential for the maintenance of GC lamination in the dentate gyrus and that GCD is the result of a local Reelin deficiency.

Potential mechanism of GABA secretion in response to the activation of GluK1-containing kainate receptors

Hippocampal GABAergic neurons are subdivided into more than 20 subtypes that are distinguished by features and functions. We have previously described the subpopulation of GABAergic neurons, which can be identified in hippocampal cell culture by the calcium response to the application of domoic acid (DoA), an agonist of kainate receptors (KARs). Here, we investigate the features of DoA-sensitive neurons and their GABA release mechanism in response to KARs activation. We demonstrate that DoA-sensitive GABAergic neurons express GluK1-containing KARs because ATPA, a selective agonist of GluK1-containing receptors, induces the calcium response exclusively in these GABAergic neurons. Our experiments also show that NASPM, previously considered a selective antagonist of calcium-permeable AMPARs, blocks calcium-permeable KARs. We established using NASPM that GluK1-containing receptors of the studied population of GABAergic neurons are calcium-permeable, and their activation is required for GABA release, at least in particular synapses. Notably, GABA release occurs even in the presence of tetrodotoxin, indicating that propagation of the depolarizing stimulus is not required for GABA release in this case. Thus, our data demonstrate that the activation of GluK1-containing calcium-permeable KARs mediates the GABA release by the studied subpopulation of GABAergic neurons.

Zinc Modulates Olfactory Bulb Kainate Receptors

Kainate receptors (KARs) are glutamate receptors with ionotropic and metabotropic activity composed of the GluK1–GluK5 subunits. We previously reported that KARs modulate excitatory and inhibitory transmission in the olfactory bulb (OB). Zinc, which is highly concentrated in the OB, also appears to modulate OB synaptic transmission via actions at other ionotropic glutamate receptors (i.e., AMPA, NMDA). However, few reports of effects of zinc on recombinant and/or native KARs exist and none have involved the OB. In the present study, we investigated the effects of exogenously applied zinc on OB KARs expressed by mitral/tufted (M/T) cells. We found that 100 µM zinc inhibits currents evoked by various combinations of KAR agonists (kainate or SYM 2081) and the AMPA receptor antagonist SYM 2206. The greatest degree of zinc-mediated inhibition was observed with coapplication of zinc with the GluK1- and GluK2-preferring agonist SYM 2081 plus SYM 2206. This finding is consistent with prior reports of zinc’s inhibitory effects on some recombinant (homomeric GluK1 and GluK2 and heteromeric GluK2/GluK4 and GluK2/GluK5) KARs, although potentiation of other (GluK3, GluK2/3) KARs has also been described. It is also of potential importance given our previously reported molecular data suggesting that OB neurons express relatively high levels of GluK1 and GluK2. Our present findings suggest that a physiologically relevant concentration of zinc modulates KARs expressed by M/T cells. As M/T cells are targets of zinc-containing olfactory sensory neurons, synaptically released zinc may influence odor information-encoding synaptic circuits in the OB via actions at KARs.

Intracerebroventricular Delivery of Human Umbilical Cord Mesenchymal Stem Cells as a Promising Therapy for Repairing the Spinal Cord Injury Induced by Kainic Acid.

Spinal cord injury (SCI) is a common pathological condition that leads to permanent or temporal loss of motor and autonomic functions. Kainic acid (KA), an agonist of kainate receptors, a type of ionotropic glutamate receptor, is widely used to induce experimental neurodegeneration models of CNS. Mesenchymal Stem Cells (MSC) therapy applied at the injured nervous tissue have emerged as a promising therapeutic treatment. Here we used a validated SCI experimental model in which an intraparenchymal injection of KA into the C5 segment of rat spinal cord induced an excitotoxic lesion. Three days later, experimental animals were treated with an intracerebroventricular injection of human umbilical cord (hUC) MSC whereas control group only received saline solution. Sensory and motor skills as well as neuronal and glial reaction of both groups were recorded. Differences in motor behavior, neuronal counting and glial responses were observed between hUC-MSC-treated and untreated rats. According to the obtained results, we suggest that hUC-MSC therapy delivered into the fourth ventricle using the intracerebroventricular via can exert a neuroprotective or neurorestorative effect on KA-injected animals.

Calcium-stimulated adenylyl cyclase subtype 1 is required for presynaptic long-term potentiation in the insular cortex of adult mice.

Recent studies indicate that presynaptic long-term potentiation in the anterior cingulate cortex may contribute to chronic pain-related anxiety. In addition to the anterior cingulate cortex, the insular cortex has also been indicated in chronic pain and its related emotional disorders. In the present study, we used a 64-channel multielectrode dish (MED64) system to record pre-long-term potentiation in the insular cortex. We showed that low-frequency stimulation paired with a GluK1-containing kainate receptor agonist induced N-methyl-D-aspartic acid receptor-independent pre-long-term potentiation in the insular cortex of wild-type mice. This form of pre-long-term potentiation was blocked in the insular cortex of adenylyl cyclase subtype 1 (AC1) knockout mice. Furthermore, a selective AC1 inhibitor NB001 blocked pre-long-term potentiation in the insular cortex with a dose-dependent manner. Taken together, our results suggest that AC1 contributes to pre-long-term potentiation in the insular cortex of adult mice and NB001 may produce anxiolytic effects by inhibiting pre-long-term potentiation in the anterior cingulate cortex and insular cortex.

Development of Cortical Pyramidal Cell and Interneuronal Dendrites: a Role for Kainate Receptor Subunits and NETO1.

During neuronal development, AMPA receptors (AMPARs) and NMDA receptors (NMDARs) are important for neuronal differentiation. Kainate receptors (KARs) are closely related to AMPARs and involved in the regulation of cortical network activity. However, their role for neurite growth and differentiation of cortical neurons is unclear. Here, we used KAR agonists and overexpression of selected KAR subunits and their auxiliary neuropilin and tolloid-like proteins, NETOs, to investigate their influence on dendritic growth and network activity in organotypic cultures of rat visual cortex. Kainate at 500nM enhanced network activity and promoted development of dendrites in layer II/III pyramidal cells, but not interneurons. GluK2 overexpression promoted dendritic growth in pyramidal cells and interneurons. GluK2 transfectants were highly active and acted as drivers for network activity. GluK1 and NETO1 specifically promoted dendritic growth of interneurons. Our study provides new insights for the roles of KARs and NETOs in the morphological and physiological development of the visual cortex.

Kainate Receptors Play a Role in Modulating Synaptic Transmission in the Olfactory Bulb

Glutamate is the neurotransmitter used at most excitatory synapses in the mammalian brain, including those in the olfactory bulb (OB). There, ionotropic glutamate receptors including N-methyl-d-aspartate receptors (NMDARs) and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) play a role in processes such as reciprocal inhibition and glomerular synchronization. Kainate receptors (KARs) represent another type of ionotropic glutamate receptor, which are composed of five (GluK1–GluK5) subunits. Whereas KARs appear to be heterogeneously expressed in the OB, evidence as to whether these KARs are functional, found at synapses, or modify synaptic transmission is limited. In the present study, coapplication of KAR agonists (kainate, SYM 2081) and AMPAR antagonists (GYKI 52466, SYM 2206) demonstrated that functional KARs are expressed by OB neurons, with a subset of receptors located at synapses. Application of kainate and the GluK1-selective agonist ATPA had modulatory effects on excitatory postsynaptic currents (EPSCs) evoked by stimulation of the olfactory nerve layer. Application of kainate and ATPA also had modulatory effects on reciprocal inhibitory postsynaptic currents (IPSCs) evoked using a protocol that evokes dendrodendritic inhibition. The latter finding suggests that KARs, with relatively slow kinetics, may play a role in circuits in which the relatively brief duration of AMPAR-mediated currents limits the role of AMPARs in synaptic transmission (e.g., reciprocal inhibition at dendrodendritic synapses). Collectively, our findings suggest that KARs, including those containing the GluK1 subunit, modulate excitatory and inhibitory transmission in the OB. These data further suggest that KARs participate in the regulation of synaptic circuits that encode odor information.

Kainate receptor deficient mast cells exhibit increased activation in vivo and in vitro.

Abstract Allergies and asthma are common ailments that are on the rise around the world, especially in young children. Mast cells play a direct role in the signs and symptoms characteristic in allergic patients. Kainate is an ionotropic member of the glutamate receptor family and is mainly studied in the brain. Recent investigations, however, have shown the presence of the Kainate Receptor (KAR) in peripheral tissues, including cells of the immune system. Our lab has previously studied the effects of KAR deficient mice and have found that these mice have reduced Th2 responses in a house dust mite extract model of allergic airway inflammation. We sought to determine if KAR deficient mast cells played a role in the phenotype observed. Paradoxically, these mast cells exhibited increased, rather than decreased mediator release in vitro by antigen–dependent degranulation. In vivo, KAR KO mice release significantly more histamine during a model of passive systemic anaphylaxis. Finally, KAR KO mice exhibit an exacerbated temperature drop in response to administered histamine. These results stand in contrast to the inhibition of the asthma model, but indicate that KAR agonist, rather than antagonist treatment of mast cells would be a treatment modality to decrease mast cell activation. Such studies are currently underway.

Targeted Deletion of Vesicular GABA Transporter from Retinal Horizontal Cells Eliminates Feedback Modulation of Photoreceptor Calcium Channels.

The cellular mechanisms underlying feedback signaling from horizontal cells to photoreceptors, which are important for the formation of receptive field surrounds of early visual neurons, remain unsettled. Mammalian horizontal cells express a complement of synaptic proteins that are necessary and sufficient for calcium-dependent exocytosis of inhibitory neurotransmitters at their contacts with photoreceptor terminals, suggesting that they are capable of releasing GABA via vesicular release. To test whether horizontal cell vesicular release is involved in feedback signaling, we perturbed inhibitory neurotransmission in these cells by targeted deletion of the vesicular GABA transporter (VGAT), the protein responsible for the uptake of inhibitory transmitter by synaptic vesicles. To manipulate horizontal cells selectively, an iCre mouse line with Cre recombinase expression controlled by connexin57 (Cx57) regulatory elements was generated. In Cx57-iCre mouse retina, only horizontal cells expressed Cre protein, and its expression occurred in all retinal regions. After crossing with a VGATflox/flox mouse line, VGAT was selectively eliminated from horizontal cells, which was confirmed immunohistochemically. Voltage-gated ion channel currents in horizontal cells of Cx57-VGAT−/− mice were the same as Cx57-VGAT+/+ controls, as were the cell responses to the ionotropic glutamate receptor agonist kainate, but the response to the GABAA receptor agonist muscimol in Cx57-VGAT−/− mice was larger. In contrast, the feedback inhibition of photoreceptor calcium channels, which in control animals is induced by horizontal cell depolarization, was completely absent in Cx57-VGAT−/− mice. The results suggest that vesicular release of GABA from horizontal cells is required for feedback inhibition of photoreceptors.

Continuous monitoring of highly reactive oxygen radicals during in vivo microdialysis

BackgroundTerephthalate (TA2−), which reacts with highly reactive oxygen species (hROS) to form the fluorophor 2-hydroxy terephthalic acid (OH-TA) with a high selectivity, has been used for determining hROS formation during in vivo microdialysis. Previously this involved collecting fractions of the microdialysate and determining the OH-TA formed after HPLC (the batch method). New methodThis work reports the development and validation of a procedure for continuously determining hROS formation during microdialysis. TA2− was added to the artificial cerebrospinal fluid (aCSF) perfusing medium to trap hROS. OH-TA formation was detected in real time with a sensitive fluorescence detector equipped with a capillary flow cell that was coupled directly to the effluent stream of the microdialysis system. ResultsThe behaviour of the system was assessed by comparison with the batch method and using a well-characterized animal model of excitotoxic damage, based on the application of high concentrations (1mM and 500μM) of the non-NMDA glutamate receptor agonist kainate (KA) to the neostriatum. Data for the evoked release of taurine were also determined in these samples. No temporal difference between hROS and taurine release could be detected. Comparison with existing method(s)The flow method had a comparable sensitivity of hROS detection to the batch method. It was simpler, cheaper and less time-consuming than the batch method. ConclusionsThis direct system is convenient and technically undemanding. It should be useful for the rapid assessment of the hROS responses to neurotoxins and other compounds in microdialysis experiments in vivo.

Minocycline does not affect long-term potentiation in the anterior cingulate cortex of normal adult mice.

It has been reported that activated microglia plays important roles in chronic pain-related sensory signaling at the spinal cord dorsal horn. Less is known about the possible contribution of microglia to cortical plasticity that has been found to be important for chronic pain. In the present study, we used a 64-channel multi-electrode array recording system to investigate the role of microglia in cortical plasticity of the anterior cingulate cortex (ACC) in normal adult mice. We found that bath application of minocycline, an inhibitor of microglial activation, had no effect on postsynaptic LTP (post-LTP) induced by theta burst stimulation in the ACC. Furthermore, presynaptic LTP (pre-LTP) induced by the combination of low-frequency stimulation with a GluK1-containing kainate receptor agonist was also not affected. The spatial distribution of post-LTP or pre-LTP among the cingulate network is also unaltered by minocycline. Our results suggest that minocycline does not affect cingulate plasticity and neurons are the major player in pain-related cortical plasticity.Electronic supplementary materialThe online version of this article (doi:10.1186/s12990-015-0025-2) contains supplementary material, which is available to authorized users.

Impaired presynaptic long-term potentiation in the anterior cingulate cortex of Fmr1 knock-out mice.

Fragile X syndrome is a common inherited form of mental impairment. Fragile X mental retardation protein (FMRP) plays important roles in the regulation of synaptic protein synthesis, and loss of FMRP leads to deficits in learning-related synaptic plasticity and behavioral disability. Previous studies mostly focus on postsynaptic long-term potentiation (LTP) in Fmr1 knock-out (KO) mice. Here, we investigate the role of FMRP in presynaptic LTP (pre-LTP) in the adult mouse anterior cingulate cortex (ACC). Low-frequency stimulation induced LTP in layer II/III pyramidal neurons under the voltage-clamp mode. Paired-pulse ratio, which is a parameter for presynaptic changes, was decreased after the low-frequency stimulation in Fmr1 wild-type (WT) mice. Cingulate pre-LTP was abolished in Fmr1 KO mice. We also used a 64-electrode array system for field EPSP recording and found that the combination of low-frequency stimulation paired with a GluK1-containing kainate receptor agonist induced NMDA receptor-independent and metabotropic glutamate receptor-dependent pre-LTP in the WT mice. This potentiation was blocked in Fmr1 KO mice. Biochemical experiments showed that Fmr1 KO mice displayed altered translocation of protein kinase A subunits in the ACC. Our results demonstrate that FMRP plays an important role in pre-LTP in the adult mouse ACC, and loss of this pre-LTP may explain some of the behavioral deficits in Fmr1 KO mice.

Rapid throughput analysis demonstrates that chemicals with distinct seizurogenic mechanisms differentially alter Ca2+ dynamics in networks formed by hippocampal neurons in culture.

Primary cultured hippocampal neurons (HN) form functional networks displaying synchronous Ca(2+) oscillations (SCOs) whose patterns influence plasticity. Whether chemicals with distinct seizurogenic mechanisms differentially alter SCO patterns was investigated using mouse HN loaded with the Ca(2+) indicator fluo-4-AM. Intracellular Ca(2+) dynamics were recorded from 96 wells simultaneously in real-time using fluorescent imaging plate reader. Although quiescent at 4 days in vitro (DIV), HN acquired distinctive SCO patterns as they matured to form extensive dendritic networks by 16 DIV. Challenge with kainate, a kainate receptor (KAR) agonist, 4-aminopyridine (4-AP), a K(+) channel blocker, or pilocarpine, a muscarinic acetylcholine receptor agonist, caused distinct changes in SCO dynamics. Kainate at <1 µM produced a rapid rise in baseline Ca(2+) (Phase I response) associated with high-frequency and low-amplitude SCOs (Phase II response), whereas SCOs were completely repressed with >1 µM kainate. KAR competitive antagonist CNQX [6-cyano-7-nitroquinoxaline-2,3-dione] (1-10 µM) normalized Ca(2+) dynamics to the prekainate pattern. Pilocarpine lacked Phase I activity but caused a sevenfold prolongation of Phase II SCOs without altering either their frequency or amplitude, an effect normalized by atropine (0.3-1 µM). 4-AP (1-30 µM) elicited a delayed Phase I response associated with persistent high-frequency, low-amplitude SCOs, and these disturbances were mitigated by pretreatment with the KCa activator SKA-31 [naphtho[1,2-d]thiazol-2-ylamine]. Consistent with its antiepileptic and neuroprotective activities, nonselective voltage-gated Na(+) and Ca(2+) channel blocker lamotrigine partially resolved kainate- and pilocarpine-induced Ca(2+) dysregulation. This rapid throughput approach can discriminate among distinct seizurogenic mechanisms that alter Ca(2+) dynamics in neuronal networks and may be useful in screening antiepileptic drug candidates.

Nitric oxide modulates the temporal properties of the glutamate response in type 4 OFF bipolar cells.

Nitric oxide (NO) is involved in retinal signal processing, but its cellular actions are only partly understood. An established source of retinal NO are NOACs, a group of nNOS-expressing amacrine cells which signal onto bipolar, other amacrine and ganglion cells in the inner plexiform layer. Here, we report that NO regulates glutamate responses in morphologically and electrophysiologically identified type 4 OFF cone bipolar cells through activation of the soluble guanylyl cyclase-cGMP-PKG pathway. The glutamate response of these cells consists of two components, a fast phasic current sensitive to kainate receptor agonists, and a secondary component with slow kinetics, inhibited by AMPA receptor antagonists. NO shortened the duration of the AMPA receptor-dependent component of the glutamate response, while the kainate receptor-dependent component remained unchanged. Application of 8-Br-cGMP mimicked this effect, while inhibition of soluble guanylate cyclase or protein kinase G prevented it, supporting a mechanism involving a cGMP signaling pathway. Notably, perfusion with a NOS-inhibitor prolonged the duration of the glutamate response, while the NO precursor L-arginine shortened it, in agreement with a modulation by endogenous NO. Furthermore, NO accelerated the response recovery during repeated stimulation of type 4 cone bipolar cells, suggesting that the temporal response properties of this OFF bipolar cell type are regulated by NO. These results reveal a novel cellular mechanism of NO signaling in the retina, and represent the first functional evidence of NO modulating OFF cone bipolar cells.

Role of GluK1 kainate receptors in seizures, epileptic discharges, and epileptogenesis.

Kainate receptors containing the GluK1 subunit have an impact on excitatory and inhibitory neurotransmission in brain regions, such as the amygdala and hippocampus, which are relevant to seizures and epilepsy. Here we used 2-amino-3-(3-hydroxy-5-tert-butylisoxazol-4-yl) propanoic acid (ATPA), a potent and selective agonist of kainate receptors that include the GluK1 subunit, in conjunction with mice deficient in GluK1 and GluK2 kainate receptor subunits to assess the role of GluK1 kainate receptors in provoking seizures and in kindling epileptogenesis. We found that systemic ATPA, acting specifically via GluK1 kainate receptors, causes locomotor arrest and forelimb extension (a unique behavioral characteristic of GluK1 activation) and induces myoclonic behavioral seizures and electrographic seizure discharges in the BLA and hippocampus. In contrast, the proconvulsant activity of systemic AMPA, kainate, and pentylenetetrazol is not mediated by GluK1 kainate receptors, and deletion of these receptors does not elevate the threshold for seizures in the 6 Hz model. ATPA also specifically activates epileptiform discharges in BLA slices in vitro via GluK1 kainate receptors. Olfactory bulb kindling developed similarly in wild-type, GluK1, and GluK2 knock-out mice, demonstrating that GluK1 kainate receptors are not required for epileptogenesis or seizure expression in this model. We conclude that selective activation of kainate receptors containing the GluK1 subunit can trigger seizures, but these receptors are not necessary for seizure generation in models commonly used to identify therapeutic agents for the treatment of epilepsy.

Fluorescent imaging reports an extracellular alkalinization induced by glutamatergic activation of isolated retinal horizontal cells

Extracellular acidification induced by retinal horizontal cells has been hypothesized to underlie lateral feedback inhibition onto vertebrate photoreceptors. To test this hypothesis, the H(+)-sensitive fluorophore 5-hexadecanoylaminofluorescein (HAF) was used to measure changes in H(+) from horizontal cells isolated from the retina of the catfish. HAF staining conditions were modified to minimize intracellular accumulation of HAF and maximize membrane-associated staining, and ratiometric fluorescent imaging of cells displaying primarily membrane-associated HAF fluorescence was conducted. Challenge of such HAF-labeled cells with glutamate or the ionotropic glutamate receptor agonist kainate produced an increase in the fluorescence ratio, consistent with an alkalinization response of +0.12 pH units and +0.23 pH units, respectively. This alkalinization was blocked by the AMPA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), the L-type calcium channel blocker nifedipine, and lanthanum. The alkalinization reported by HAF was consistent with extracellular alkalinizations detected in previous studies using self-referencing H(+)-selective microelectrodes. The spatial distribution of the kainate-induced changes in extracellular H(+) was also examined. An overall global alkalinization around the cell was observed, with no obvious signs of discrete centers of acidification. Taken together, these data argue against the hypothesis that glutamatergic-induced efflux of protons from horizontal cells mediates lateral feedback inhibition in the outer retina.