RESUMEN
In retinitis pigmentosa (RP), rod and cone photoreceptors degenerate, depriving downstream neurons of light-sensitive input, leading to vision impairment or blindness. Although downstream neurons survive, some undergo morphological and physiological remodeling. Bipolar cells (BCs) link photoreceptors, which sense light, to retinal ganglion cells (RGCs), which send information to the brain. While photoreceptor loss disrupts input synapses to BCs, whether BC output synapses remodel has remained unknown. Here we report that synaptic output from BCs plummets in RP mouse models of both sexes owing to loss of voltage-gated Ca2+ channels. Remodeling reduces the reliability of synaptic output to repeated optogenetic stimuli, causing RGC firing to fail at high-stimulus frequencies. Fortunately, functional remodeling of BCs can be reversed by inhibiting the retinoic acid receptor (RAR). RAR inhibitors targeted to BCs present a new therapeutic opportunity for mitigating detrimental effects of remodeling on signals initiated either by surviving photoreceptors or by vision-restoring tools.
Asunto(s)
Células Bipolares de la Retina , Sinapsis , Tretinoina , Animales , Células Bipolares de la Retina/efectos de los fármacos , Células Bipolares de la Retina/fisiología , Ratones , Tretinoina/farmacología , Masculino , Femenino , Sinapsis/efectos de los fármacos , Sinapsis/fisiología , Retinitis Pigmentosa/fisiopatología , Retinitis Pigmentosa/genética , Degeneración Retiniana/fisiopatología , Ratones Endogámicos C57BL , Ratones Transgénicos , Células Ganglionares de la Retina/fisiología , Células Ganglionares de la Retina/efectos de los fármacosRESUMEN
Retinal ganglion cells (RGCs) receive glutamatergic input from bipolar cells through NMDA- and AMPA-type glutamate receptors. Both GluA2-containing, Ca(2+)-impermeable AMPA receptors (CI-AMPARs) and GluA2-lacking, Ca(2+)-permeable AMPA receptors (CP-AMPARs) contribute to light-evoked responses in ON RGCs; however, specific roles for each subtype are not well understood. Here, we present evidence that light intensity determines the subtype of AMPAR that is activated during the synaptic response in ON RGCs. Using current voltage analysis of the EPSC we show that light intensities near RGC threshold, intensities that travel through the well described primary rod pathway, evoke synaptic currents that are preferentially mediated by CP-AMPARs. Synaptic responses evoked by spontaneous release of transmitter from bipolar cell terminals also preferentially activate CP-AMPARs. Conversely, higher light intensities, most likely carried by secondary rod pathways, activate CI-AMPARs. The same pattern of CP-AMPAR and CI-AMPAR activation was observed in mice containing only functional rods, suggesting that the recruitment of CI-AMPARs at higher light intensity does not require cone stimulation. When glutamate spillover was induced by blocking transporters with TBOA, both the near threshold and spontaneous EPSCs contained a significant CI-AMPAR component. We propose that CI-AMPARs are activated by "spillover" of synaptic glutamate only during bright illumination, or when glutamate uptake is blocked. Glutamate may spill over to more distant sites at the same synapse, or perhaps as far as neighboring synapses. Together, our data suggest that the spatial organization of AMPARs at ON RGCs synapses allows for selective, intensity-dependent activation of AMPARs with distinct subunit composition.
Asunto(s)
Receptores AMPA/metabolismo , Células Ganglionares de la Retina/metabolismo , Animales , Potenciales Postsinápticos Excitadores/fisiología , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Técnicas de Placa-Clamp , Estimulación LuminosaRESUMEN
ON bipolar cells are critical for the function of the ON pathway in the visual system. They express a metabotropic glutamate receptor (mGluR6) that, when activated, couples to the G(o) class of G protein. The channel that is primarily responsible for the synaptic response has been recently identified as the transient receptor potential cation channel subfamily M member 1 (TRPM1); TRPM1 is negatively coupled to the mGluR6/Go cascade such that activation of the cascade results in closure of the channel. Light indirectly opens TRPM1 by reducing transmitter release from presynaptic photoreceptors, resulting in a decrease in mGluR6 activation. Conversely, in the dark, binding of synaptic glutamate to mGluR6 inhibits TRPM1 current. Closure of TRPM1 by G-protein activation in the dark is a critical step in the process of ON bipolar cell signal transduction, but the precise pathway linking these two events is not understood. To address this question, we measured TRPM1 activity in retinal bipolar cells, in human ependymal melanocytes (HEMs) that endogenously express TRPM1, and in HEK293 cells transfected with TRPM1. Dialysis of the Gßγ subunit dimer, but not Gα(o), closed TRPM1 channels in every cell type that we tested. In addition, activation of an endogenous G-protein-coupled receptor pathway in HEK293 cells that releases Gßγ without activating Go protein also closed TRPM1 channels. These results suggest a model in which the Gßγ dimer that is released as a result of the dissociation from Gα(o) upon activation of mGluR6 closes the TRPM1 channel, perhaps via a direct interaction.
Asunto(s)
Subunidades alfa de la Proteína de Unión al GTP Gi-Go/metabolismo , Subunidades beta de la Proteína de Unión al GTP/metabolismo , Subunidades gamma de la Proteína de Unión al GTP/metabolismo , Canales Catiónicos TRPM/fisiología , Aminoácidos/farmacología , Animales , Capsaicina/análogos & derivados , Capsaicina/farmacología , Células Cultivadas , Antagonistas de Aminoácidos Excitadores/farmacología , Subunidades beta de la Proteína de Unión al GTP/química , Subunidades gamma de la Proteína de Unión al GTP/química , Ácido Glutámico/metabolismo , Células HEK293 , Humanos , Luz , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/efectos de la radiación , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Unión Proteica/efectos de los fármacos , Unión Proteica/efectos de la radiación , Multimerización de Proteína , Receptores de Glutamato Metabotrópico/antagonistas & inhibidores , Receptores de Glutamato Metabotrópico/metabolismo , Células Bipolares de la Retina/efectos de los fármacos , Células Bipolares de la Retina/metabolismo , Células Bipolares de la Retina/fisiología , Canales Catiónicos TRPM/genética , Canales Catiónicos TRPM/metabolismo , Xantenos/farmacologíaRESUMEN
Retinal ganglion cells receive inputs from multiple bipolar cells which must be integrated before a decision to fire is made. Theoretical studies have provided clues about how this integration is accomplished but have not directly determined the rules regulating summation of closely timed inputs along single or multiple dendrites. Here we have examined dendritic summation of multiple inputs along On ganglion cell dendrites in whole mount rat retina. We activated inputs at targeted locations by uncaging glutamate sequentially to generate apparent motion along On ganglion cell dendrites in whole mount retina. Summation was directional and dependent13 on input sequence. Input moving away from the soma (centrifugal) resulted in supralinear summation, while activation sequences moving toward the soma (centripetal) were linear. Enhanced summation for centrifugal activation was robust as it was also observed in cultured retinal ganglion cells. This directional summation was dependent on hyperpolarization activated cyclic nucleotide-gated (HCN) channels as blockade with ZD7288 eliminated directionality. A computational model confirms that activation of HCN channels can override a preference for centripetal summation expected from cell anatomy. This type of direction selectivity could play a role in coding movement similar to the axial selectivity seen in locust ganglion cells which detect looming stimuli. More generally, these results suggest that non-directional retinal ganglion cells can discriminate between input sequences independent of the retina network.
Asunto(s)
Modelos Neurológicos , Sumación de Potenciales Postsinápticos/fisiología , Células Ganglionares de la Retina/fisiología , Animales , Biología Computacional , Canales Catiónicos Regulados por Nucleótidos Cíclicos/metabolismo , Dendritas/fisiología , Glutamatos/metabolismo , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización , Procesos Fotoquímicos , Canales de Potasio/metabolismo , Pirimidinas , Ratas , Ratas Sprague-DawleyRESUMEN
On retinal ganglion cells (RGCs) transmit light encoded information to the brain and receive excitatory input from On cone bipolar cells (CBPs). The synaptic CBP input onto On RGCs is mediated by AMPA-type glutamate receptors (AMPARs) that include both those lacking a GluA2 subunit, and are therefore permeable to Ca(2+), and those that possess at least one GluA2 subunit and are Ca(2+)-impermeable. We have previously demonstrated in electrophysiological studies that periods of low synaptic activity, brought about by housing animals in darkness, enhance the proportion of GluA2-lacking AMPARs at the On CBP-On RGC synapse by mobilizing surface GluA2 containing receptors into a receptor pool that rapidly cycles in and out of the membrane. AMPAR cycling induction by reduced synaptic activity takes several hours. This delay suggests that changes in expression of proteins which regulate AMPAR trafficking may mediate the altered mobility of GluA2 AMPARs in RGCs. In this study, we test the hypothesis that AMPAR trafficking proteins couple synaptic activity to AMPAR cycling in RGCs. Immunocytochemical and biochemical analyses confirmed that darkness decreases surface GluA2 in RGCs and changed the expression levels of three proteins associated with GluA2 trafficking. GRIP was decreased, while PICK1 and Arc were increased. Knockdown of GRIP with siRNA elevated constitutive AMPAR cycling, mimicking effects of reduced synaptic activity, while knockdown of PICK1 and Arc blocked increases in constitutive GluA2 trafficking. Our results support a role for correlated, activity-driven changes in multiple AMPAR trafficking proteins that modulate GluA2 cycling which can in turn affect synaptic AMPAR composition in RGCs.
Asunto(s)
Receptores AMPA/metabolismo , Células Ganglionares de la Retina/metabolismo , Animales , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/metabolismo , Péptidos y Proteínas de Señalización Intercelular , Péptidos y Proteínas de Señalización Intracelular , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Unión Proteica , Transporte de Proteínas , Ratas , Ratas Sprague-Dawley , Receptores AMPA/genética , Células Ganglionares de la Retina/fisiología , Membranas Sinápticas/metabolismo , Transmisión SinápticaRESUMEN
The ciliary body, located at the junction of the choroid and iris, is crucial in the development of the embryonic eye. Notch2 signalling, Wnt signalling, transforming growth factor ß (TGF-ß) signalling, and Pax6 signalling are critical for coordinating the ciliary body formation. These signalling pathways are coordinated with each other and participate in the ciliary body development, ensuring the precise formation and optimal functioning of the eye structure. Although rare, ciliary body hypoplasia, ciliary tumours, and genetic-related iritis indicate the intricate nature of ciliary body development. Given the ciliary body's important biological significance and potential medical relevance, we aim to provide a comprehensive overview of the developmental molecular mechanisms governing ciliary body formation and function. Here, we focus on the intricate signalling pathways governing ciliary body development and corresponding genetic ciliary diseases.
RESUMEN
Bipolar cells of the retina carry visual information from photoreceptors in the outer retina to retinal ganglion cells (RGCs) in the inner retina. Bipolar cells express L-type voltage-gated Ca2+ channels at the synaptic terminal, but generally lack other types of channels capable of regenerative activity. As a result, the flow of information from outer to inner retina along bipolar cell processes is generally passive in nature, with no opportunity for signal boost or amplification along the way. Here we report the surprising discovery that blocking voltage-gated Na+ channels profoundly reduces the synaptic output of one class of bipolar cell, the type 6 ON bipolar cell (CBC6), despite the fact that the CBC6 itself does not express voltage-gated Na+ channels. Instead, CBC6 borrows voltage-gated Na+ channels from its neighbor, the inhibitory AII amacrine cell, with whom it is connected via an electrical synapse. Thus, an inhibitory neuron aids in amplification of an excitatory signal as it moves through the retina, ensuring that small changes in the membrane potential of bipolar cells are reliably passed onto downstream RGCs.
RESUMEN
In the retina, light onset hyperpolarizes photoreceptors and depolarizes ON bipolar cells at the sign inverting photoreceptor-ON bipolar cell synapse. Transmission at this synapse is mediated by a signaling cascade comprised of mGluR6, a G-protein containing G(αo), and the cation channel TRP melastatin 1 (TRPM1). This system is thought to be common to both the rod- and ON-cone-driven pathways, which control vision under scotopic and photopic conditions, respectively. In this study, we present evidence that the rod pathway is uniquely susceptible to modulation by PKCα at the rod-rod bipolar cell synapse. Decreased production of DAG (an activator of PKC) by inhibition of PIP2 (phosphatidylinositol-4,5-bisphosphate) hydrolysis caused depression of the TRPM1 current. Conversely, addition of a DAG analog, 2-acetyl-1-oleoyl-sn-glycerol (OAG), potentiated the current in rod bipolar cells but not in ON-cone bipolar cells. The potentiating effects of OAG were absent both in mutant mice that lack PKCα expression and in wild-type mice in which enzymatic activity of PKCα was pharmacologically inhibited. In addition, we found that, like other members of the TRPM subfamily, TRPM1 current is susceptible to voltage-independent inhibition by intracellular magnesium, and that modulation by PKCα relieves this inhibition, as the potentiating effects of OAG are absent in low intracellular magnesium. We conclude that activation of PKCα initiates a modulatory mechanism at the rod-rod bipolar cell synapse whose function is to reduce inhibition of the TRPM1 current by magnesium, thereby increasing the gain of transmission at this synapse.
Asunto(s)
Magnesio/fisiología , Proteína Quinasa C-alfa/fisiología , Células Bipolares de la Retina/fisiología , Células Fotorreceptoras Retinianas Conos/fisiología , Células Fotorreceptoras Retinianas Bastones/fisiología , Sinapsis/fisiología , Canales Catiónicos TRPM/fisiología , Animales , Diglicéridos/farmacología , Inhibidores Enzimáticos/farmacología , Femenino , Técnicas In Vitro , Magnesio/metabolismo , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/genética , Potenciales de la Membrana/fisiología , Ratones , Ratones Mutantes , Fosfatidilinositol 4,5-Difosfato/antagonistas & inhibidores , Fosfatidilinositol 4,5-Difosfato/metabolismo , Fosfolipasa D/antagonistas & inhibidores , Proteína Quinasa C-alfa/antagonistas & inhibidores , Proteína Quinasa C-alfa/biosíntesis , Proteína Quinasa C-alfa/genética , Células Bipolares de la Retina/efectos de los fármacos , Células Bipolares de la Retina/metabolismo , Células Fotorreceptoras Retinianas Conos/efectos de los fármacos , Células Fotorreceptoras Retinianas Conos/metabolismo , Células Fotorreceptoras Retinianas Bastones/efectos de los fármacos , Células Fotorreceptoras Retinianas Bastones/metabolismo , Sinapsis/metabolismo , Canales Catiónicos TRPM/antagonistas & inhibidoresRESUMEN
Expression of channels to specific neuronal sites can critically impact their function and regulation. Currently, the molecular mechanisms underlying this targeting and intracellular trafficking of transient receptor potential (TRP) channels remain poorly understood, and identifying proteins involved in these processes will provide insight into underlying mechanisms. Vision is dependent on the normal function of retinal depolarizing bipolar cells (DBCs), which couple a metabotropic glutamate receptor 6 to the TRP melastatin 1 (TRPM1) channel to transmit signals from photoreceptors. We report that the extracellular membrane-attached protein nyctalopin is required for the normal expression of TRPM1 on the dendrites of DBCs in mus musculus. Biochemical and genetic data indicate that nyctalopin and TRPM1 interact directly, suggesting that nyctalopin is acting as an accessory TRP channel subunit critical for proper channel localization to the synapse.
Asunto(s)
Dendritas/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Proteínas/metabolismo , Retina/citología , Células Bipolares de la Retina/citología , Canales Catiónicos TRPM/metabolismo , Aminoácidos/farmacología , Animales , Proteínas Bacterianas/genética , Biofisica , Línea Celular Transformada , Estimulación Eléctrica , Antagonistas de Aminoácidos Excitadores/farmacología , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Humanos , Técnicas In Vitro , Proteínas Repetidas Ricas en Leucina , Proteínas Luminiscentes/genética , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Modelos Biológicos , Técnicas de Placa-Clamp , Proteínas/genética , Proteoglicanos/genética , Proteoglicanos/metabolismo , Retina/crecimiento & desarrollo , Canales Catiónicos TRPM/deficiencia , Transfección/métodos , Xantenos/farmacologíaRESUMEN
ON bipolar cells invert the sign of light responses from hyperpolarizing to depolarizing before passing them on to ganglion cells. Light responses are generated when a cation channel, recently identified as Trpm1, opens. The amplitude of the light response rapidly decays due to desensitization of Trpm1 current. The role of Trpm1 desensitization in shaping light responses both in bipolar and downstream ganglion cells has not been well characterized. Here we show that two parameters, the amount and the rate of recovery from desensitization, depend on the strength of the presynaptic stimulus. Stimuli that activate less than 20% of the maximum Trpm1 current did not promote any detectable desensitization, even for prolonged periods. Beyond this threshold there was a linear relationship between the amount of desensitization and the fractional Trpm1 current. In response to stimuli that open all available channels, desensitization reduced the response to approximately 40% of the peak, with a time constant of 1 s, and recovery was slow, with a time constant of more than 20 s. In dye-filled bipolar cells classified as transient or sustained using morphological criteria, there were no significant differences in Trpm1 desensitization parameters. Trpm1 activation evoked robust EPSCs in ganglion cells, and removal of Trpm1 desensitization strongly augmented a sustained component of the ganglion cell EPSC irrespective of whether ganglion cells were of the ON or ON/OFF type. We conclude that Trpm1 desensitization impacts the kinetics of ganglion cell EPSCs, but does not underlie the sustained/transient dichotomy of neurons in the ON pathway.
Asunto(s)
Células Bipolares de la Retina/metabolismo , Canales Catiónicos TRPM/metabolismo , Ambystoma , Animales , Calcio/metabolismo , Canales Iónicos/metabolismo , Luz , Retina/metabolismo , Retina/fisiología , Células Ganglionares de la Retina/metabolismo , Transducción de Señal , Sinapsis/metabolismo , Potenciales Sinápticos/fisiologíaRESUMEN
Mutations in TRPM1 are found in humans with an autosomal recessive form of complete congenital stationary night blindness (cCSNB). The Trpm1(-/-) mouse has been an important animal model for this condition. Here we report a new mouse mutant, tvrm27, identified in a chemical mutagenesis screen. Genetic mapping of the no b-wave electroretinogram (ERG) phenotype of tvrm27 localized the mutation to a chromosomal region that included Trpm1. Complementation testing with Trpm1(-/-) mice confirmed a mutation in Trpm1. Sequencing identified a nucleotide change in exon 23, converting a highly conserved alanine within the pore domain to threonine (p.A1068T). Consistent with prior studies of Trpm1(-/-) mice, no anatomical changes were noted in the Trpm1(tvrm27/tvrm27) retina. The Trpm1(tvrm27/tvrm27) phenotype is distinguished from that of Trpm1(-/-) by the retention of TRPM1 expression on the dendritic tips of depolarizing bipolar cells (DBCs). While ERG b-wave amplitudes of Trpm1(+/-) heterozygotes are comparable to wild type, those of Trpm1(+/tvrm27) mice are reduced by 32%. A similar reduction in the response of Trpm1(+/tvrm27) DBCs to LY341495 or capsaicin is evident in whole cell recordings. These data indicate that the p.A1068T mutant TRPM1 acts as a dominant negative with respect to TRPM1 channel function. Furthermore, these data indicate that the number of functional TRPM1 channels at the DBC dendritic tips is a key factor in defining DBC response amplitude. The Trpm1(tvrm27/tvrm27) mutant will be useful for elucidating the role of TRPM1 in DBC signal transduction, for determining how Trpm1 mutations impact central visual processing, and for evaluating experimental therapies for cCSNB.
Asunto(s)
Mutación Puntual , Células Bipolares de la Retina/fisiología , Canales Catiónicos TRPM/genética , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/fisiología , Secuencia de Aminoácidos , Aminoácidos/farmacología , Animales , Capsaicina/farmacología , Mapeo Cromosómico , Cromosomas de los Mamíferos/genética , Dendritas/fisiología , Modelos Animales de Enfermedad , Exones , Enfermedades Hereditarias del Ojo/genética , Enfermedades Genéticas Ligadas al Cromosoma X/genética , Heterocigoto , Ratones , Ratones Transgénicos , Datos de Secuencia Molecular , Mutación Missense , Miopía/genética , Ceguera Nocturna/genética , Retina/patología , Retina/fisiología , Análisis de Secuencia de ADN , Canales Catiónicos TRPM/metabolismo , Treonina/genética , Xantenos/farmacologíaRESUMEN
On bipolar cells are connected to photoreceptors via a sign-inverting synapse. At this synapse, glutamate binds to a metabotropic receptor which couples to the closure of a cation-selective transduction channel. The molecular identity of both the receptor and the G protein are known, but the identity of the transduction channel has remained elusive. Here, we show that the transduction channel in mouse rod bipolar cells, a subtype of On bipolar cell, is likely to be a member of the TRP family of channels. To evoke a transduction current, the metabotropic receptor antagonist LY341495 was applied to the dendrites of cells that were bathed in a solution containing the mGluR6 agonists L-AP4 or glutamate. The transduction current was suppressed by ruthenium red and the TRPV1 antagonists capsazepine and SB-366791. Furthermore, focal application of the TRPV1 agonists capsaicin and anandamide evoked a transduction-like current. The capsaicin-evoked and endogenous transduction current displayed prominent outward rectification, a property of the TRPV1 channel. To test the possibility that the transduction channel is TRPV1, we measured rod bipolar cell function in the TRPV1(-/-) mouse. The ERG b-wave, a measure of On bipolar cell function, as well as the transduction current and the response to TRPV1 agonists were normal, arguing against a role for TRPV1. However, ERG measurements from mice lacking TRPM1 receptors, another TRP channel implicated in retinal function, revealed the absence of a b-wave. Our results suggest that a TRP-like channel, possibly TRPM1, is essential for synaptic function in On bipolar cells.
Asunto(s)
Células Bipolares de la Retina/fisiología , Transmisión Sináptica/fisiología , Canales de Potencial de Receptor Transitorio/metabolismo , Aminoácidos/farmacología , Anilidas/farmacología , Animales , Ácidos Araquidónicos/farmacología , Capsaicina/análogos & derivados , Capsaicina/farmacología , Cinamatos/farmacología , Endocannabinoides , Antagonistas de Aminoácidos Excitadores/farmacología , Ácido Glutámico/metabolismo , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Alcamidas Poliinsaturadas/farmacología , Propionatos/farmacología , Receptores de Glutamato Metabotrópico/agonistas , Receptores de Glutamato Metabotrópico/metabolismo , Rojo de Rutenio/metabolismo , Transmisión Sináptica/efectos de los fármacos , Canales Catiónicos TRPM/agonistas , Canales Catiónicos TRPM/antagonistas & inhibidores , Canales Catiónicos TRPM/metabolismo , Canales Catiónicos TRPV/agonistas , Canales Catiónicos TRPV/antagonistas & inhibidores , Canales Catiónicos TRPV/metabolismo , Canales de Potencial de Receptor Transitorio/agonistas , Canales de Potencial de Receptor Transitorio/antagonistas & inhibidores , Xantenos/farmacologíaRESUMEN
AMPA-type glutamate receptors in the CNS are normally impermeable to Ca2+, but the aberrant expression of Ca2+-permeable AMPA receptors (CP-AMPARs) occurs in pathological conditions such as ischemia or epilepsy, or degenerative diseases such as ALS. Here, we show that select populations of retinal ganglion cells (RGCs) similarly express high levels of CP-AMPARs in a mouse model of glaucoma. CP-AMPAR expression increased dramatically in both On sustained alpha and Off transient alpha RGCs, and this increase was prevented by genomic editing of the GluA2 subunit. On sustained alpha RGCs with elevated CP-AMPAR levels displayed profound synaptic depression, which was reduced by selectively blocking CP-AMPARs, buffering Ca2+ with BAPTA, or with the CB1 antagonist AM251, suggesting that depression was mediated by a retrograde transmitter which might be triggered by the influx of Ca2+ through CP-AMPARs. Thus, glaucoma may alter the composition of AMPARs and depress excitatory synaptic input in select populations of RGCs.
RESUMEN
Vision impairment and blindness in humans are most frequently caused by the degeneration and loss of photoreceptor cells in the outer retina, as is the case for age-related macular degeneration, retinitis pigmentosa, retinal detachment and many other diseases. While inner retinal neurons survive degeneration, they undergo fundamental pathophysiological changes, collectively known as "remodeling." Inner retinal remodeling downstream to photoreceptor death occurs across mammalian retinas from mice to humans, independently of the cause of degeneration. It results in pervasive spontaneous hyperactivity and membrane hyperpermeability in retinal ganglion cells, which funnel all retinal signals to the brain. Remodeling reduces light detection in vision-impaired patients and precludes meaningful vision restoration in blind individuals. In this review, we summarize current hypotheses proposed to explain remodeling and their potential medical significance highlighting the important role played by retinoic acid and its receptor.
RESUMEN
Horizontal cells (HCs) form reciprocal synapses with rod and cone photoreceptors, an arrangement that underlies lateral inhibition in the retina. HCs send negative and positive feedback signals to photoreceptors, but how HCs initiate these signals remains unclear. Unfortunately, because HCs have no unique neurotransmitter receptors, there are no pharmacological treatments for perturbing membrane potential specifically in HCs. Here we use transgenic zebrafish whose HCs express alien receptors, enabling cell-type-specific control by cognate alien agonists. To depolarize HCs, we used the Phe-Met-Arg-Phe-amide (FMRFamide)-gated Na+ channel (FaNaC) activated by the invertebrate neuropeptide FMRFamide. To hyperpolarize HCs we used a pharmacologically selective actuator module (PSAM)-glycine receptor (GlyR), an engineered Cl- selective channel activated by a synthetic agonist. Expression of FaNaC or PSAM-GlyR was restricted to HCs with the cell-type selective promoter for connexin-55.5. We assessed HC-feedback control of photoreceptor synapses in three ways. First, we measured presynaptic exocytosis from photoreceptor terminals using the fluorescent dye FM1-43. Second, we measured the electroretinogram (ERG) b-wave, a signal generated by postsynaptic responses. Third, we used Ca2+ imaging in retinal ganglion cells (RGCs) expressing the Ca2+ indicator GCaMP6. Addition of FMRFamide significantly decreased FM1-43 destaining in darkness, whereas the addition of PSAM-GlyR significantly increased it. However, both agonists decreased the light-elicited ERG b-wave and eliminated surround inhibition of the Ca2+ response of RGCs. Taken together, our findings show that chemogenetic tools can selectively manipulate negative feedback from HCs, providing a platform for understanding its mechanism and helping to elucidate its functional roles in visual information processing at a succession of downstream stages.
Asunto(s)
Células Horizontales de la Retina , Pez Cebra , Animales , Potenciales de la Membrana , Retina , Células Fotorreceptoras Retinianas ConosRESUMEN
At conventional synapses, invasion of an action potential into the presynaptic terminal produces a rapid Ca(2+) influx and ultimately the release of synaptic vesicles. However, retinal rod bipolar cells (RBCs) generally do not produce action potentials, and the rate of depolarization of the axon terminal is instead governed by the rate of rise of the light response, which can be quite slow. Using paired whole-cell recordings, we measured the behaviour of the RBC-AII amacrine cell synapse while simulating light-induced depolarizations either by slowly ramping the RBC voltage or by depolarizing the RBC with the mGluR6 receptor antagonist (R,S)-alpha-cyclopropyl-4-phosphonophenylglycine (CPPG). Both voltage ramps and CPPG evoked slow activation of presynaptic Ca(2+) currents and severely attenuated the early, transient component of the AII EPSC compared with voltage steps. We also found that the duration of the transient component was limited in time, and this limitation could not be explained by vesicle depletion, inhibitory feedback, or proton inhibition. Limiting the duration of the fast transient insures the availability of readily releasable vesicles to support a second, sustained component of release. The mGluR6 pathway modulator cGMP sped the rate of RBC depolarization in response to puffs of CPPG and consequently potentiated the transient component of the EPSC at the expense of the sustained component. We conclude that the rod bipolar cell is capable of both transient and sustained signalling, and modulation of the mGluR6 pathway by cGMP allows the RBC to switch between these two time courses of transmitter release.
Asunto(s)
Células Amacrinas/metabolismo , Células Fotorreceptoras Retinianas Bastones/metabolismo , Transmisión Sináptica , Visión Ocular , Células Amacrinas/efectos de los fármacos , Animales , Señalización del Calcio , GMP Cíclico/metabolismo , Potenciales Postsinápticos Excitadores , Glicina/análogos & derivados , Glicina/farmacología , Técnicas In Vitro , Cinética , Ratones , Ratones Endogámicos C57BL , Técnicas de Placa-Clamp , Terminales Presinápticos/metabolismo , Receptores de Glutamato Metabotrópico/antagonistas & inhibidores , Receptores de Glutamato Metabotrópico/metabolismo , Células Fotorreceptoras Retinianas Bastones/efectos de los fármacos , Transmisión Sináptica/efectos de los fármacos , Vesículas Sinápticas/metabolismo , Visión Ocular/efectos de los fármacosRESUMEN
Synaptic transmission from photoreceptors to all types of ON bipolar cells is primarily mediated by the mGluR6 receptor. This receptor, which is apparently expressed uniquely in the nervous system by ON bipolar cells, couples negatively to a nonselective cation channel. This arrangement results in a sign reversal at photoreceptor/ON bipolar cell synapse, which is necessary in order to establish parallel ON and OFF pathways in the retina. The synapse is an important target for second messenger molecules that are known to modulate synaptic transmission elsewhere in the nervous system, second messengers that act on a time scale ranging from milliseconds to minutes. This review focuses on two of these molecules, Ca2+ and cGMP, summarizing our current knowledge of how they modulate gain at the photoreceptor/ON bipolar cell synapse, as well as their proposed sites of action within the mGluR6 cascade. The implications of plasticity at this synapse for retinal function will also be examined.
Asunto(s)
Células Bipolares de la Retina/metabolismo , Calcio/fisiología , GMP Cíclico/fisiología , Humanos , Células Fotorreceptoras de Vertebrados/fisiología , Receptores de Glutamato Metabotrópico/metabolismo , Transmisión Sináptica/fisiología , Visión Ocular/fisiologíaRESUMEN
In this review, we summarize studies investigating the types and distribution of voltage- and calcium-gated ion channels in the different classes of retinal neurons: rods, cones, horizontal cells, bipolar cells, amacrine cells, interplexiform cells, and ganglion cells. We discuss differences among cell subtypes within these major cell classes, as well as differences among species, and consider how different ion channels shape the responses of different neurons. For example, even though second-order bipolar and horizontal cells do not typically generate fast sodium-dependent action potentials, many of these cells nevertheless possess fast sodium currents that can enhance their kinetic response capabilities. Ca2+ channel activity can also shape response kinetics as well as regulating synaptic release. The L-type Ca2+ channel subtype, CaV1.4, expressed in photoreceptor cells exhibits specific properties matching the particular needs of these cells such as limited inactivation which allows sustained channel activity and maintained synaptic release in darkness. The particular properties of K+ and Cl- channels in different retinal neurons shape resting membrane potentials, response kinetics and spiking behavior. A remaining challenge is to characterize the specific distributions of ion channels in the more than 100 individual cell types that have been identified in the retina and to describe how these particular ion channels sculpt neuronal responses to assist in the processing of visual information by the retina.
Asunto(s)
Canales Iónicos/fisiología , Neuronas Retinianas/fisiología , Células Amacrinas/fisiología , Animales , Humanos , Células Fotorreceptoras Retinianas Conos/fisiología , Células Ganglionares de la Retina/fisiología , Células Fotorreceptoras Retinianas Bastones/fisiología , Transmisión Sináptica/fisiologíaRESUMEN
Progressive degeneration of retinal ganglion cells (RGCs) will cause a blinding disease. Most of the study is focusing on the RGCs itself. In this study, we demonstrate a decline of the presynaptic rod bipolar cells (RBCs) response precedes RGCs loss and a decrease of protein kinase Cα (PKCα) protein expression in RBCs dendrites, using whole-cell voltage-clamp, electroretinography (ERG) measurements, immunostaining and co-immunoprecipitation. We present evidence showing that N-methyl D-aspartate receptor subtype 2B (NR2B)/protein interacting with C kinase 1 (PICK1)-dependent degradation of PKCα protein in RBCs contributes to RBCs functional loss. Mechanistically, NR2B forms a complex with PKCα and PICK1 to promote the degradation of PKCα in a phosphorylation- and proteasome-dependent manner. Similar deficits in PKCα expression and response sensitivity were observed in acute ocular hypertension and optic never crush models. In conclusion, we find that three separate experimental models of neurodegeneration, often used to specifically target RGCs, disrupt RBCs function prior to the loss of RGCs. Our findings provide useful information for developing new diagnostic tools and treatments for retinal ganglion cells degeneration disease.
Asunto(s)
Neurotoxinas/toxicidad , Células Bipolares de la Retina/patología , Células Ganglionares de la Retina/patología , Células Fotorreceptoras Retinianas Bastones/patología , Animales , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Proteínas de Ciclo Celular/metabolismo , Dendritas/efectos de los fármacos , Dendritas/metabolismo , Modelos Animales de Enfermedad , Electrorretinografía , Glaucoma/patología , Células HEK293 , Humanos , Masculino , Ratones Endogámicos C57BL , N-Metilaspartato/toxicidad , Degeneración Nerviosa/patología , Fosforilación/efectos de los fármacos , Complejo de la Endopetidasa Proteasomal/metabolismo , Unión Proteica/efectos de los fármacos , Proteína Quinasa C-alfa/metabolismo , Proteolisis/efectos de los fármacos , Receptores de N-Metil-D-Aspartato/metabolismo , Células Bipolares de la Retina/efectos de los fármacos , Células Fotorreceptoras Retinianas Conos/efectos de los fármacos , Células Fotorreceptoras Retinianas Conos/patología , Células Ganglionares de la Retina/efectos de los fármacos , Células Fotorreceptoras Retinianas Bastones/efectos de los fármacos , Ubiquitina/metabolismoRESUMEN
At many excitatory synapses, AMPA-type receptors (AMPARs) are not statically situated in the membrane, but undergo continuous rounds of endocytosis and exocytosis, referred to as rapid cycling. AMPAR cycling is believed to play a role in certain forms of synaptic plasticity, but the link between cycling and synaptic function is not well understood. We have previously demonstrated that AMPARs cycle in neurons of the inner retina, including amacrine and ganglion cells, and that cycling is inhibited by synaptic activity. Recording from cultured neurons and ON ganglion cells in the flat-mount retina, we now show that rapid cycling is primarily, perhaps exclusively, restricted to AMPARs that contain the GluR2 subunit, and that cycle is confined to extrasynaptic receptors. We also demonstrate a form of plasticity at the ON bipolar cell-ON ganglion cell synapse, whereby synaptic quiescence drives a change in the composition of AMPARs from predominantly GluR2-containing to GluR2-lacking. Finally, we provide evidence linking synaptic receptor composition and cycling, showing that disruption of cycling leads increases the number of GluR2-containing receptors in the ON bipolar-ON ganglion cell synapse. We propose that cycling lowers the number of GluR2-containing receptors at the surface and, consequently, within the synapse. After increased levels of synaptic activity, cycling ceases, and all GluR2-containing receptors are free to go to the surface, where they can be delivered to synapses. Our results suggest that by regulating the cycling of AMPARs, ambient light can modulate the composition of synaptic receptors in ON ganglion cells.