Altered Forebrain Functional Connectivity and Neurotransmission in a Kinase-Inactive Met Mouse Model of Autism.

MET, the gene encoding the tyrosine kinase receptor for hepatocyte growth factor, is a susceptibility gene for autism spectrum disorder (ASD). Genetically altered mice with a kinase-inactive Met offer a potential model for understanding neural circuit organization changes in autism. Here, we focus on the somatosensory thalamocortical circuitry because distinct somatosensory sensitivity phenotypes accompany ASD, and this system plays a major role in sensorimotor and social behaviors in mice. We employed resting-state functional magnetic resonance imaging and in vivo high-resolution proton MR spectroscopy to examine neuronal connectivity and neurotransmission of wild-type, heterozygous Met-Emx1, and fully inactive homozygous Met-Emx1 mice. Met-Emx1 brains showed impaired maturation of large-scale somatosensory network connectivity when compared with wild-type controls. Significant sex × genotype interaction in both network features and glutamate/gamma-aminobutyric acid (GABA) balance was observed. Female Met-Emx1 brains showed significant connectivity and glutamate/GABA balance changes in the somatosensory thalamocortical system when compared with wild-type brains. The glutamate/GABA ratio in the thalamus was correlated with the connectivity between the somatosensory cortex and the thalamus in heterozygous Met-Emx1 female brains. The findings support the hypothesis that aberrant functioning of the somatosensory thalamocortical system is at the core of the conspicuous somatosensory behavioral phenotypes observed in Met-Emx1 mice.

Insulin-Independent GABAA Receptor-Mediated Response in the Barrel Cortex of Mice with Impaired Met Activity.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder caused by genetic variants, susceptibility alleles, and environmental perturbations. The autism associated geneMETtyrosine kinase has been implicated in many behavioral domains and endophenotypes of autism, including abnormal neural signaling in human sensory cortex. We investigated somatosensory thalamocortical synaptic communication in mice deficient in Met activity in cortical excitatory neurons to gain insights into aberrant somatosensation characteristic of ASD. The ratio of excitation to inhibition is dramatically increased due to decreased postsynaptic GABAAreceptor-mediated inhibition in the trigeminal thalamocortical pathway of mice lacking active Met in the cerebral cortex. Furthermore, in contrast to wild-type mice, insulin failed to increase GABAAreceptor-mediated response in the barrel cortex of mice with compromised Met signaling. Thus, lacking insulin effects may be a risk factor in ASD pathogenesis. SIGNIFICANCE STATEMENT: A proposed common cause of neurodevelopmental disorders is an imbalance in excitatory neural transmission, provided by the glutamatergic neurons, and the inhibitory signals from the GABAergic interneurons. Many genes associated with autism spectrum disorders impair synaptic transmission in the expected cell type. Previously, inactivation of the autism-associated Met tyrosine kinase receptor in GABAergic interneurons led to decreased inhibition. In thus report, decreased Met signaling in glutamatergic neurons had no effect on excitation, but decimated inhibition. Further experiments indicate that loss of Met activity downregulates GABAAreceptors on glutamatergic neurons in an insulin independent manner. These data provide a new mechanism for the loss of inhibition and subsequent abnormal excitation/inhibition balance and potential molecular candidates for treatment or prevention.

Enhancement of postsynaptic GABAA and extrasynaptic NMDA receptor-mediated responses in the barrel cortex of Mecp2-null mice.

Rett syndrome (RTT) is a neurodevelopmental disorder that results from mutations in the X-linked gene for methyl-CpG-binding protein 2 (MECP2). The underlying cellular mechanism for the sensory deficits in patients with RTT is largely unknown. This study used the Bird mouse model of RTT to investigate sensory thalamocortical synaptic transmission in the barrel cortex of Mecp2-null mice. Electrophysiological results showed an excitation/inhibition imbalance, biased toward inhibition, due to an increase in efficacy of postsynaptic GABAA receptors rather than alterations in inhibitory network and presynaptic release properties. Enhanced inhibition impaired the transmission of tonic sensory signals from the thalamus to the somatosensory cortex. Previous morphological studies showed an upregulation of NMDA receptors in the neocortex of both RTT patients and Mecp2-null mice at early ages [Blue ME, Naidu S, Johnston MV. Ann Neurol 45: 541-545, 1999; Blue ME, Kaufmann WE, Bressler J, Eyring C, O'Driscoll C, Naidu S, Johnston MV. Anat Rec (Hoboken) 294: 1624-1634, 2011]. Although AMPA and NMDA receptor-mediated excitatory synaptic transmission was not altered in the barrel cortex of Mecp2-null mice, extrasynaptic NMDA receptor-mediated responses increased markedly. These responses were blocked by memantine, suggesting that extrasynaptic NMDA receptors play an important role in the pathogenesis of RTT. The results suggest that enhancement of postsynaptic GABAA and extrasynaptic NMDA receptor-mediated responses may underlie impaired somatosensation and that pharmacological blockade of extrasynaptic NMDA receptors may have therapeutic value for RTT.

Sensory Activity-Dependent and Sensory Activity-Independent Properties of the Developing Rodent Trigeminal Principal Nucleus.

The whisker-sensory trigeminal central pathway of rodents is an established model for studies of activity-dependent neural plasticity. The first relay station of the pathway is the trigeminal principal nucleus (PrV), the ventral part of which receives sensory inputs mainly from the infraorbital branch of the maxillary trigeminal nerve (ION). Whisker-sensory afferents play an important role in the development of the morphological and physiological properties of PrV neurons. In neonates, deafferentation by ION transection leads to the disruption of whisker-related neural patterns (barrelettes) and cell death within a specific time window (critical period), as revealed by morphological studies. Whisker-sensory inputs control synaptic elimination, postsynaptic AMPA receptor trafficking, astrocyte-mediated synaptogenesis, and receptive-field characteristics of PrV cells, without a postnatal critical period. Sensory activity-dependent synaptic plasticity requires the activation of NMDA receptors and involves the participation of glia. However, the basic physiological properties of PrV neurons, such as cell type-specific ion channels, presynaptic terminal function, postsynaptic NMDA receptor subunit composition, and formation of the inhibitory circuitry, are independent of sensory inputs. Therefore, the first relay station of the whisker sensation is largely mature-like and functional at birth. Delineation of activity-dependent and activity-independent features of the postnatal PrV is important for understanding the development and functional characteristics of downstream trigeminal stations in the thalamus and neocortex. This mini review focuses on such features of the developing rodent PrV.

Thalamic NMDA receptor function is necessary for patterning of the thalamocortical somatosensory map and for sensorimotor behaviors.

NMDARs play a major role in patterning of topographic sensory maps in the brain. Genetic knock-out of the essential subunit of NMDARs in excitatory cortical neurons prevents whisker-specific neural pattern formation in the barrel cortex. To determine the role of NMDARs en route to the cortex, we generated sensory thalamus-specific NR1 (Grin1)-null mice (ThNR1KO). A multipronged approach, using histology, electrophysiology, optical imaging, and behavioral testing revealed that, in these mice, whisker patterns develop in the trigeminal brainstem but do not develop in the somatosensory thalamus. Subsequently, there is no barrel formation in the neocortex yet a partial afferent patterning develops. Whisker stimulation evokes weak cortical activity and presynaptic neurotransmitter release probability is also affected. We found several behavioral deficits in tasks, ranging from sensorimotor to social and cognitive. Collectively, these results show that thalamic NMDARs play a critical role in the patterning of the somatosensory thalamic and cortical maps and their impairment may lead to pronounced behavioral defects.

Neonatal infraorbital nerve crush-induced CNS synaptic plasticity and functional recovery.

Infraorbital nerve (ION) transection in neonatal rats leads to disruption of whisker-specific neural patterns (barrelettes), conversion of functional synapses into silent synapses, and reactive gliosis in the brain stem trigeminal principal nucleus (PrV). Here we tested the hypothesis that neonatal peripheral nerve crush injuries permit better functional recovery of associated central nervous system (CNS) synaptic circuitry compared with nerve transection. We developed an in vitro whisker pad-trigeminal ganglion (TG)-brain stem preparation in neonatal rats and tested functional recovery in the PrV following ION crush. Intracellular recordings revealed that 68% of TG cells innervate the whisker pad. We used the proportion of whisker pad-innervating TG cells as an index of ION function. The ION function was blocked by ∼64%, immediately after mechanical crush, then it recovered beginning after 3 days postinjury and was complete by 7 days. We used this reversible nerve-injury model to study peripheral nerve injury-induced CNS synaptic plasticity. In the PrV, the incidence of silent synapses increased to ∼3.5 times of control value by 2-3 days postinjury and decreased to control levels by 5-7 days postinjury. Peripheral nerve injury-induced reaction of astrocytes and microglia in the PrV was also reversible. Neonatal ION crush disrupted barrelette formation, and functional recovery was not accompanied by de novo barrelette formation, most likely due to occurrence of recovery postcritical period (P3) for pattern formation. Our results suggest that nerve crush is more permissive for successful regeneration and reconnection (collectively referred to as "recovery" here) of the sensory inputs between the periphery and the brain stem.

Functional significance of cortical NMDA receptors in somatosensory information processing.

N-methyl-d-aspartate receptor (NMDAR)-mediated activity is required for whisker-related neural patterning in the rodent brain. Deletion of the essential NMDAR subunit NR1 gene in excitatory cortical neurons prevents whisker-specific barrel formation and impairs thalamocortical afferent patterning. We used electrophysiological and voltage-sensitive dye imaging methods to assess synaptic and sensory evoked cortical activity and immunohistochemistry to examine immediate early gene expression following whisker stimulation in cortex-specific NR1 knockout (CxNR1KO) mice. In mutant mice, layer IV neurons lacked NMDAR-mediated excitatory postsynaptic currents, and temporal summation of excitatory postsynaptic potentials (EPSPs) was impaired. Barrel neurons showed both phasic and tonic responses to whisker deflection. The averaged tonic response in CxNR1KO mice was significantly less than that in control mice due to impaired EPSP temporal summation. Electrophysiological estimation of the number of thalamic neurons innervating single barrel neurons indicated a significant increase in CxNR1KO mice. Similarly, voltage-sensitive dye optical signals in response to whisker stimulation were widespread. Immediate early gene expression following whisker stimulation also showed a diffuse expression pattern in the CxNR1KO cortex compared with whisker-specific expression patterns in controls. Thus, when NMDAR function is impaired, spatial discrimination of whisker inputs is severely compromised, and sensory stimulation evokes diffuse, topographically misaligned activity in the barrel cortex.

Peripheral nerve damage does not alter release properties of developing central trigeminal afferents.

The infraorbital branch of the trigeminal nerve (ION) is essential in whisker-specific neural patterning ("barrelettes") in the principal nucleus of the trigeminal nerve (PrV). The barrelettes are formed by the ION terminal arbors, somata, and dendrites of the PrV cells; they are abolished after neonatal damage to the ION. Physiological studies show that disruption of the barrelettes is accompanied by conversion of functional synapses into silent synapses in the PrV. In this study, we used whole cell recordings with a paired-pulse stimulation protocol and MK-801 blocking rate to estimate the presynaptic release probability (Pr) of ION central trigeminal afferent terminals in the PrV. We investigated Pr during postnatal development, following neonatal ION damage, and determined whether conversion of functional synapses into silent synapses after peripheral denervation results from changes in Pr. The paired-pulse ratio (PPR) was quite variable ranging from 40% (paired-pulse depression) to 175% (paired-pulse facilitation). The results from paired-pulse protocol were confirmed by MK-801 blocking rate experiments. The nonuniform PPRs did not show target cell specificity and developmental regulation. The distribution of PPRs fit nicely to Gaussian function with a peak at ∼ 100%. In addition, neonatal ION transections did not alter the distribution pattern of PPR in their central terminals, suggesting that the conversion from functional synapses into silent synapses in the peripherally denervated PrV is not caused by changes in the Pr.

Astrocytes promote peripheral nerve injury-induced reactive synaptogenesis in the neonatal CNS.

Neonatal damage to the trigeminal nerve leads to "reactive synaptogenesis" in the brain stem sensory trigeminal nuclei. In vitro models of brain injury-induced synaptogenesis have implicated an important role for astrocytes. In this study we tested the role of astrocyte function in reactive synaptogenesis in the trigeminal principal nucleus (PrV) of neonatal rats following unilateral transection of the infraorbital (IO) branch of the trigeminal nerve. We used electrophysiological multiple input index analysis (MII) to estimate the number of central trigeminal afferent fibers that converge onto single barrelette neurons. In the developing PrV, about 30% of afferent connections are eliminated within 2 postnatal weeks. After neonatal IO nerve damage, multiple trigeminal inputs (2.7 times that of the normal inputs) converge on single barrelette cells within 3-5 days; they remain stable up to the second postnatal week. Astrocyte proliferation and upregulation of astrocyte-specific proteins (GFAP and ALDH1L1) accompany reactive synaptogenesis in the IO nerve projection zone of the PrV. Pharmacological blockade of astrocyte function, purinergic receptors, and thrombospondins significantly reduced or eliminated reactive synaptogenesis without changing the MII in the intact PrV. GFAP immunohistochemistry further supported these electrophysiological results. We conclude that immature astrocytes, purinergic receptors, and thrombospondins play an important role in reactive synaptogenesis in the peripherally deafferented neonatal PrV.

LTD and LTP at the developing retinogeniculate synapse.

The purpose of the present study was to determine whether retinal activity can support long-term changes in synaptic strength in the developing dorsal lateral geniculate nucleus (LGN) of thalamus. To test for this we made use of a rodent in vitro explant preparation in which retinal afferents and the intrinsic circuitry of the LGN remain intact. We repetitively stimulated the optic tract with a tetanus protocol that approximated the temporal features of spontaneous retinal waves. We found the amplitude of extracellular field potentials evoked by retinal stimulation changed significantly after tetanus and that the polarity of these alterations was related to postnatal age. At a time when substantial pruning of retinal connections occurs (postnatal day 1 [P1] to P14), high-frequency stimulation led to an immediate and long-term depression (LTD). However, at times when pruning wanes and adult-like patterns of connectivity are stabilizing (P16 to P30), the identical form of stimulation produced a modest form of potentiation (long-term potentiation [LTP]). The LTD was unaffected by the bath application of gamma-aminobutyric acid type A and N-methyl-D-aspartate receptor antagonists. However, both LTD and LTP were blocked by L-type Ca(2+)-channel antagonists. Thus the Ca(2+) influx associated with L-type channel activation mediates the induction of synaptic plasticity and may signal the pruning and subsequent stabilization of developing retinogeniculate connections.

Conversion of functional synapses into silent synapses in the trigeminal brainstem after neonatal peripheral nerve transection.

One of the major consequences of neonatal infraorbital nerve damage is irreversible morphological reorganization in the principal sensory nucleus (PrV) of the trigeminal nerve in the brainstem. We used the voltage-clamp technique to study synaptic transmission in the normal and the denervated PrV of neonatal rats in an in vitro brainstem preparation. Most of the synapses in the PrV are already functional at birth. Three days after peripheral deafferentation, functional synapses become silent, lacking AMPA receptor-mediated currents. Without sensory inputs from the whiskers, silent synapses persist through the second postnatal week, indicating that the maintenance of AMPA receptor function depends on sensory inputs. High-frequency (50 Hz) electrical stimulation of the afferent pathway, which mimics sensory input, restores synaptic function, whereas low-frequency (1 Hz) stimulation has no effect. Application of glycine, which promotes AMPA receptor exocytosis, also restores synaptic function. Therefore, normal synaptic function in the developing PrV requires incoming activity via sensory afferents and/or enhanced AMPA receptor exocytosis. Sensory deprivation most likely results in AMPA receptor endocytosis and/or lateral diffusion to the extrasynaptic membrane.

Insulin receptor sensitization restores neocortical excitation/inhibition balance in a mouse model of autism.

Background: Met receptor tyrosine kinase regulates neurogenesis, differentiation, migration, connectivity, and synaptic plasticity. The human Met gene has been identified as a prominent risk factor for autism spectrum disorder (ASD). Met gene-altered mice serve as useful models for mechanistic studies of ASD. Inactivation of Met in excitatory cortical neurons in mice (Emx1cre/Metflox mice) yields a phenotype in which significantly decreased GABAA receptor-mediated inhibition shifts the excitation/inhibition (E/I) balance toward excitation in the somatosensory cortex. Further, unlike that seen in wild-type mice, insulin does not increase inhibition in the mutant cortex, suggesting that one of the consequences of kinase inactive Met gene could be desensitization of insulin receptors. To test this hypothesis, we investigated the effects of insulin receptor sensitizer, pioglitazone, on inhibition in the somatosensory thalamocortical circuitry. Methods: We used whole-cell patch clamp electrophysiology and analyzed excitatory and inhibitory responses of cortical layer IV excitatory cells following stimulation of their thalamic input in thalamocortical pathway intact brain slices. We applied insulin alone and insulin + a thiazolidinedione, pioglitazone (PIO), to test the effects of sensitizing insulin receptors on inhibitory responses mediated by GABAA receptors in the somatosensory cortex of Emx1cre/Metflox mice. Results: In WT brain slices, application of insulin together with PIO did not enhance the effect of insulin alone. In contrast, PIO application induced a much larger inhibition than that of insulin alone in Met-defective cortex. Thus, insulin resistance of GABAA receptor-mediated response in Met mutant mice may result from desensitized insulin receptors. Conclusions: Sporadic clinical studies reported improved behavioral symptoms in children with autism following PIO treatment. We show that PIO can aid in normalization of the E/I balance in the primary somatosensory cortex, a potential physiological mechanism underlying the positive effects of PIO treatment.

NMDA receptor-dependent regulation of axonal and dendritic branching.

In the rodent trigeminal principal nucleus (PrV), trigeminal afferent terminals and postsynaptic cells form discrete modules ("barrelettes") that replicate the patterned array of whiskers and sinus hairs on the snout. Barrelette neurons of the PrV relay whisker-specific patterns to the contralateral thalamus and, subsequently, to the primary somatosensory barrel cortex. Genetic impairment of NMDA receptor (NMDAR) function blocks development of barrelettes in the PrV. Underlying cellular and functional defects are not known. Here, we examined morphological differentiation of whisker afferents, dendritic differentiation of barrelette cells, and their electrophysiological properties in mice with genetic perturbations of the essential subunit NR1 of NMDARs. We show that in NR1 gene knock-down (KD) and knock-out mice, whisker afferents begin their embryonic development normally but, over time, fail to segregate into patches, and instead they develop exuberant terminal arbors spanning most of the PrV. Postnatal NR1KD barrelette cells, with significantly reduced NMDA currents, retain their membrane and synaptic properties but develop longer dendrites with no orientation preference. These results indicate that NMDARs regulate growth of presynaptic terminal arbors and postsynaptic dendritic branching, thereby leading to consolidation of synapses and patterning of presynaptic and postsynaptic elements.

Neonatal sensory nerve injury-induced synaptic plasticity in the trigeminal principal sensory nucleus.

Sensory deprivation studies in neonatal mammals, such as monocular eye closure, whisker trimming, and chemical blockade of the olfactory epithelium have revealed the importance of sensory inputs in brain wiring during distinct critical periods. But very few studies have paid attention to the effects of neonatal peripheral sensory nerve damage on synaptic wiring of the central nervous system (CNS) circuits. Peripheral somatosensory nerves differ from other special sensory afferents in that they are more prone to crush or severance because of their locations in the body. Unlike the visual and auditory afferents, these nerves show regenerative capabilities after damage. Uniquely, damage to a somatosensory peripheral nerve does not only block activity incoming from the sensory receptors but also mediates injury-induced neuro- and glial chemical signals to the brain through the uninjured central axons of the primary sensory neurons. These chemical signals can have both far more and longer lasting effects than sensory blockade alone. Here we review studies which focus on the consequences of neonatal peripheral sensory nerve damage in the principal sensory nucleus of the brainstem trigeminal complex.

Dependence of Retinogeniculate Transmission on Membrane Voltage in the Cat.

The mammalian lateral geniculate nucleus seems organized to gate or control the gain of retinogeniculate transmission, the result of which is then relayed to the visual cortex. We have performed in vivo intracellular studies of retinogeniculate transmission along these retino-geniculo-cortical pathways in cats by recording the retinally evoked excitatory postsynaptic potential (EPSP) in geniculate neurons. In cats, these pathways are organized into two parallel and functionally distinct channels, the X and Y pathways. We found that nearly all geniculate X cells display a fairly conventional voltage dependency for their retinally evoked EPSPs, because the amplitudes of these EPSPs decrease fairly linearly with membrane depolarization as the EPSP reversal potential is approached. Rare X cells and all Y cells, however, show an unconventional response: over a wide range of membrane potentials, their EPSP amplitudes increase with membrane depolarization. This increase does not result from alterations in neuronal input resistance and instead seems due to changes in synaptic conductance. The underlying cause of this voltage dependency remains to be determined. None the less, it does afford an interesting means by which retinogeniculate transmission can be gated, since non-retinal inputs (e.g. corticogeniculate axons) that can control a relay Y cell's membrane potential can also modulate the cell's EPSP amplitude.

Structural and functional composition of the developing retinogeniculate pathway in the mouse.

The advent of transgenic mice has made the developing retinogeniculate pathway a model system for targeting potential mechanisms that underlie the refinement of sensory connections. However, a detailed characterization of the form and function of this pathway is lacking. Here we use a variety of anatomical and electrophysiological techniques to delineate the structural and functional changes occurring in the lateral geniculate nucleus (LGN) of dorsal thalamus of the C57/BL6 mouse. During the first two postnatal weeks there is an age-related recession in the amount of terminal space occupied by retinal axons arising from the two eyes. During the first postnatal week, crossed and uncrossed axons show substantial overlap throughout most of the LGN. Between the first and second week retinal arbors show significant pruning, so that by the time of natural eye opening (P12-14) segregation is complete and retinal projections are organized into distinct eye-specific domains. During this time of rapid anatomical rearrangement, LGN cells could be readily distinguished using immunocytochemical markers that stain for NMDA receptors, GABA receptors, L-type Ca2+ channels, and the neurofilament protein SMI-32. Moreover, the membrane properties and synaptic responses of developing LGN cells are remarkably stable and resemble those of mature neurons. However, there are some notable developmental changes in synaptic connectivity. At early ages, LGN cells are binocularly responsive and receive input from as many as 11 different retinal ganglion cells. Optic tract stimulation also evokes plateau-like depolarizations that are mediated by the activation of L-type Ca2+ channels. As retinal inputs from the two eyes segregate into nonoverlapping territories, there is a loss of binocular responsiveness, a decrease in retinal convergence, and a reduction in the incidence of plateau potentials. These data serve as a working framework for the assessment of phenotypes of genetically altered strains as well as provide some insight as to the molecular mechanisms underlying the refinement of retinogeniculate connections.

Properties of LTD and LTP of retinocollicular synaptic transmission in the developing rat superior colliculus.

The developing retinocollicular pathway undergoes synaptic refinement in order to form the precise retinotopic pattern seen in adults. To study the mechanisms which underlie refinement, we investigated long-term changes in retinocollicular transmission in rats aged P0-P25. Field potentials (FPs) in the superior colliculus (SC) were evoked by stimulation of optic tract fibers in an in vitro isolated brainstem preparation. High intensity stimulation induced long-term depression (LTD) in the SC after both low (1000 stimuli at 1 Hz) and higher (1000 stimuli at 50 Hz) frequency stimulation. The induction of LTD was independent of activation of NMDA and GABA(A) receptors, because D-APV (100 microM) and bicuculline (10 microM) did not block LTD. Induction of LTD was dependent upon activation of L-type Ca(2+) channels as 10 microM nitrendipine, an L-type Ca(2+) channel blocker, significantly decreased the magnitude of LTD. LTD was down-regulated during development. LTD magnitude was greatest in rats aged P0-P9 and significantly less in rats aged P10-P25. Long-term potentiation (LTP) was induced by low intensity stimulation and only after high frequency tetanus (1000 stimuli at 50 Hz). LTP was NMDA receptor dependent because d-APV (100 microM) completely abolished it. LTP induction was also blocked by the L-type Ca2+ channel blocker nitrendipine. The magnitude of LTP first increased with age, being significantly greater at P7-P13 than at P0-3 and then decreased at P23-25. In summary, both LTD and LTP are present during retinocollicular pathway refinement, but have different transmitter and ionic mechanisms and time courses of expression.

L-type calcium channel-mediated plateau potentials in barrelette cells during structural plasticity.

Development and maintenance of whisker-specific patterns along the rodent trigeminal pathway depends on an intact sensory periphery during the sensitive/critical period in development. Barrelette cells of the brain stem trigeminal nuclei are the first set of neurons to develop whisker-specific patterns. Those in the principal sensory nucleus (PrV) relay these patterns to the ventrobasal thalamus, and consequently, to the somatosensory cortex. Thus PrV barrelette cells are among the first group of central neurons susceptible to the effects of peripheral damage. Previously we showed that membrane properties of barrelette cells are distinct as early as postnatal day 1 (PND 1) and remain unchanged following peripheral denervation in newborn rat pups (Lo and Erzurumlu 2001). In the present study, we investigated the changes in synaptic transmission. In barrelette cells of normal PND 1 rats, weak stimulation of the trigeminal tract (TrV) that was subthreshold for inducing Na(+) spikes evoked an excitatory postsynaptic potential-inhibitory postsynaptic potential (EPSP-IPSP) sequence that was similar to the responses seen in older rats (Lo et al. 1999). Infraorbital nerve transection at birth did not alter excitatory and inhibitory synaptic connections of the barrelette cells. These observations suggested that local neuronal circuits are already established in PrV at birth and remain intact after deafferentation. Strong stimulation of the TrV induced a sustained depolarization (plateau potential) in denervated but not in normal barrelette neurons. The plateau potential was distinct from the EPSP-IPSP sequence by 1) a sustained (>80 ms) depolarization above -40 mV; 2) a slow decline slope (<0.1 mV/ms); 3) partially or totally inactivated Na(+) spikes on the plateau; and 4) a termination by a steep decay (>1 mV/ms) to a hyperpolarizing membrane level. The plateau potential was mediated by L-type Ca(2+) channels and triggered by a N-methyl-D-aspartate (NMDA) receptor-mediated EPSP. gamma-aminobutyric acid-A (GABA(A)) receptor-mediated IPSP dynamically regulated the latency and duration of the plateau potential. These results indicate that after neonatal peripheral damage, central trigeminal inputs cause a large and long-lasting Ca(2+) influx through L-type Ca(2+) channels in barrelette neurons. Increased Ca(2+) entry may play a key role in injury-induced structural remodeling, and/or transsynaptic cell death.

Synaptic mechanisms regulating the activation of a Ca(2+)-mediated plateau potential in developing relay cells of the LGN.

Using intracellular recordings in an isolated (in vitro) rat brain stem preparation, we examined the synaptic responses of developing relay neurons in the dorsal lateral geniculate nucleus (LGN). In newborn rats, strong stimulation of the optic tract (OT) evoked excitatory postsynaptic potentials (EPSPs) that gave rise to a sustained (300-1,300 ms), slow-decaying (<0.01 mV/s), depolarization (25-40 mV). Riding atop this response was a train of spikes of variable amplitude. We refer to this synaptically evoked event as a plateau potential. Pharmacology experiments indicate the plateau potential was mediated by the activation of high-threshold L-type Ca(2+) channels. Synaptic activation of the plateau potential relied on N-methyl-D-aspartate (NMDA) receptor-mediated activity and the spatial and/or temporal summation of retinally evoked EPSPs. Inhibitory postsynaptic responses (IPSPs) did not prevent the expression of the plateau potential. However, GABA(A) receptor activity modulated the intensity of optic tract stimulation needed to evoke the plateau potential, while GABA(B) receptor activity affected its duration. Expression of the plateau potential was developmentally regulated, showing a much higher incidence at P1-2 (90%) than at P19-20 (1%). This was in part due to the fact that developing relay cells show a greater degree of spatial summation than their mature counterparts, receiving input from as many as 7-12 retinal ganglion cells. Early spontaneous retinal activity is also likely to trigger the plateau potential. Repetitive stimulation of optic tract in a manner that approximated the high-frequency discharge of retinal ganglion cells led to a massive temporal summation of EPSPs and the activation of a sustained depolarization (>1 min) that was blocked by L-type Ca(2+) channel antagonists. These age-related changes in Ca(2+) signaling may contribute to the activity-dependent refinement of retinogeniculate connections.

Nature of inhibitory postsynaptic activity in developing relay cells of the lateral geniculate nucleus.

Using intracellular recordings in an isolated (in vitro) brain stem preparation, we examined the inhibitory postsynaptic responses of developing neurons in the dorsal lateral geniculate nucleus (LGN) of the rat. As early as postnatal day (P) 1-2, 31% of all excitatory postsynaptic (EPSP) activity evoked by electrical stimulation of the optic tract was followed by inhibitory postsynaptic potentials (IPSPs). By P5, 98% of all retinally evoked EPSPs were followed by IPSP activity. During the first postnatal week, IPSPs were mediated largely by GABA(A) receptors. Additional GABA(B)-mediated IPSPs emerged at P3-4 but were not prevalent until after the first postnatal week. Experiments involving the separate stimulation of each optic nerve indicated that developing LGN cells were binocularly innervated. At P11-14, it was common to evoke EPSP/IPSP pairs by stimulating either the contralateral or ipsilateral optic nerve. During the third postnatal week, binocular excitatory responses were encountered far less frequently. However, a number of cells still maintained a binocular inhibitory response. These results provide insight about the ontogeny and nature of postsynaptic inhibitory activity in the LGN during the period of retinogeniculate axon segregation.