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Interneuron-based cell transplantation is a powerful method to modify network function in a variety of neurological disorders, including epilepsy. Whether new interneurons integrate into native neural networks in a subtype-specific manner is not well understood, and the therapeutic mechanisms underlying interneuron-based cell therapy, including the role of synaptic inhibition, are debated. In this study, we tested subtype-specific integration of transplanted interneurons using acute cortical brain slices and visualized patch-clamp recordings to measure excitatory synaptic inputs, intrinsic properties, and inhibitory synaptic outputs. Fluorescently labeled progenitor cells from the embryonic medial ganglionic eminence (MGE) were used for transplantation. At 5 wk after transplantation, MGE-derived parvalbumin-positive (PV+) interneurons received excitatory synaptic inputs, exhibited mature interneuron firing properties, and made functional synaptic inhibitory connections to native pyramidal cells that were comparable to those of native PV+ interneurons. These findings demonstrate that MGE-derived PV+ interneurons functionally integrate into subtype-appropriate physiological niches within host networks following transplantation.

The postnatal functions of the Dlx1&2 transcription factors in cortical interneurons (CINs) are unknown. Here, using conditional Dlx1, Dlx2, and Dlx1&2 knockouts (CKOs), we defined their roles in specific CINs. The CKOs had dendritic, synaptic, and survival defects, affecting even PV+ CINs. We provide evidence that DLX2 directly drives Gad1, Gad2, and Vgat expression, and show that mutants had reduced mIPSC amplitude. In addition, the mutants formed fewer GABAergic synapses on excitatory neurons and had reduced mIPSC frequency. Furthermore, Dlx1/2 CKO had hypoplastic dendrites, fewer excitatory synapses, and reduced excitatory input. We provide evidence that some of these phenotypes were due to reduced expression of GRIN2B (a subunit of the NMDA receptor), a high confidence Autism gene. Thus, Dlx1&2 coordinate key components of CIN postnatal development by promoting their excitability, inhibitory output, and survival.

Abnormal GABAergic interneuron density, and imbalance of excitatory versus inhibitory tone, is thought to result in epilepsy, neurodevelopmental disorders, and psychiatric disease. Recent studies indicate that interneuron cortical density is determined primarily by the size of the precursor pool inthe embryonic telencephalon. However, factors essential for regulating interneuron allocation from telencephalic multipotent precursors are poorly understood. Here we report that Olig1 represses production of GABAergic interneurons throughout the mouse brain. Olig1 deletion in mutant mice results in ectopic expression and upregulation of Dlx1/2 genes in the ventral medial ganglionic eminences and adjacent regions of the septum, resulting in an ~30% increase in adult cortical interneuron numbers. We show that Olig1 directly represses the. Dlx1/2 I12b intergenic enhancer and that Dlx1/2 functions genetically downstream of Olig1. These findings establish Olig1 as an essential repressor of. Dlx1/2 and interneuron production in developing mammalian brain. © 2014 Elsevier Inc.

Significance We describe homeostatic plasticity of both excitatory synaptic transmission and intrinsic properties of CA1 pyramidal neurons in distal-less homeobox 1 ( Dlx1 −/− ) mice, a genetic model of postdevelopment interneuron cell death. Loss of synaptic inhibition and compensation by excitation led to enhanced potential for long-term potentiation (LTP) and altered neural oscillations. This shows that homeostatic compensation may rebalance inhibition and excitation but cannot fully normalize neural function. Transplantation of interneuron progenitor cells restored inhibitory synaptic transmission to WT levels and induced a reversal of homeostatic changes to excitation and excitability. LTP and gamma oscillations were also normalized after integration of transplanted interneurons. These data indicate that homeostatic plasticity functions in vivo to balance activity based on inhibitory tone.

Dlx transcription factors are important in the differentiation of GABAergic interneurons. In mice lacking Dlx1, early steps in interneuron development appear normal. Beginning at ∼1 mo of age, primarily dendrite-innervating interneuron subtypes begin to undergo apoptosis in Dlx1−/− mice; this is accompanied by a reduction in GABAergic transmission and late-onset epilepsy. The reported reduction of synaptic inhibition is greater than might be expected given that interneuron loss is relatively modest in Dlx1−/− mice. Here we report that voltage-clamp recordings of CA1 interneurons in hippocampal slices prepared from Dlx1−/− animals older than postnatal day 30 (>P30) revealed a significant reduction in excitatory postsynaptic current (EPSC) amplitude. No changes in EPSCs onto interneurons were observed in cells recorded from younger animals (P9–12). Current-clamp recordings from interneurons at these early postnatal ages showed that interneurons in Dlx1−/− mutants were immature and more excitable, although membrane properties normalized by P30. Terminal deoxynucleotidyl transferase dUTP-mediated nick-end labeling, caspase-3, and NeuN staining did not reveal frank cell damage or loss in area CA3 of hippocampal sections from adult Dlx1−/− mice. Delayed interneuron maturation may lead to interneuron hyperexcitability, followed by a compensatory reduction in the strength of excitatory transmission onto interneurons. This reduced excitation onto surviving interneurons, coupled with the loss of a significant fraction of GABAergic inputs to excitatory neurons starting at P30, may underlie cortical dysrhythmia and seizures previously observed in adult Dlx1−/− mice.

Mature nucleus magnocellularis (NM) neurons, the avian homolog of bushy cells of the mammalian anteroventral cochlear nucleus, maintain high [Cl−]iand depolarize in response to GABA. Depolarizing GABAergic postsynaptic potentials (GPSPs) activate both the synaptic conductance and large outward currents, which, when coupled together, inhibit spikes via shunting and spike threshold accommodation. We studied the maturation of the synaptic and voltage-dependent components of inhibition in embryonic NM neurons using whole-cell and gramicidin-perforated patch-clamp techniques to measure Cl−reversal potential, GABAergic synaptic responses, and voltage-dependent outward currents. We found that GABA enhanced excitability in immature NM neurons, undergoing a switch to inhibitory between embryonic day 14 (E14) and E18. Low-voltage-activated Kv1-type (dendrotoxin-I sensitive) K+currents increased in amplitude between E14 and E18, whereas Cl−reversal potential and synaptic conductances remained relatively stable during this period. GABA was rendered inhibitory because of this increase in low-voltage activated outward currents. GPSPs summed with other inputs to increase spike probability at E14. GPSPs shunted spikes at E18, but blocking Kv1 channels transformed this inhibition to excitation, similar to E14 neurons. Subthreshold depolarizing current steps, designed to activate outward currents similar to depolarizing GPSPs, enhanced excitability at E14 but inhibited spiking in E18 neurons. Blocking Kv1 channels reversed this effect, rendering current steps excitatory. We present the novel finding that the developmental transition of GABAergic processing from increasing neuronal excitability to inhibiting spiking can depend on changes in the expression of voltage-gated channels rather than on a change in Cl−reversal potential.

Ephrins and Eph receptors are a family of molecules that have been implicated in many developmental processes including neuronal network formation, guidance of cell migration, and axonal pathfinding. These molecules exhibit the ability to send bidirectional signals following ligand–receptor interactions resulting from cell–cell contacts. Gene-targeted knockout mice of B-class ephrins and Eph receptors have been shown to display phenotypic responses that correlate with anatomical defects. For example, disruption of the EphB2 receptor leads to defects of the vestibular system, including pathfinding abnormalities in efferent axons and reduced endolymph production. Such developmental distortions lead to deficiencies in ionic homeostasis and repetitive circling behaviors. The present study demonstrates that B-class ephrins and Eph receptors are expressed in cochlear tissues, suggesting that they may play some role in auditory function. To determine whether ephrins and Eph receptors have a functional role in the peripheral auditory system, distortion-product otoacoustic emission (DPOAE) levels, collected across a broad frequency range, were compared between groups of mice expressing different Eph receptor genotypes. In particular, EphB1 and EphB3 receptor knockout mice exhibited significantly diminished DPOAE levels as compared to wild-type littermates, indicating that these specific Eph receptors are necessary for normal cochlear function.