Highlights • Mice devoid of thalamic adenylyl cyclase 1 (AC1) are generated. • Thalamic AC1 is required for cortical barrel formation. • Thalamic AC1 is required for refinement of thalamocortical ...axons. • Cortical calcium-stimulated adenylyl cyclases are dispensable for barrel formation.
The neuronal circuits mediating the sedative action of diazepam are unknown. Although the motor-depressant action of diazepam is suppressed in alpha1(H101R) homozygous knockin mice expressing ...diazepam-insensitive alpha1-GABA(A) receptors, global alpha1-knockout mice show greater motor sedation with diazepam. To clarify this paradox, attributed to compensatory up-regulation of the alpha2 and alpha3 subunits, and to further identify the neuronal circuits supporting diazepam-induced sedation, we generated Emx1-cre-recombinase-mediated conditional mutant mice, selectively lacking the alpha1 subunit (forebrain-specific alpha1(-/-)) or expressing either a single wild-type (H) or a single point-mutated (R) alpha1 allele (forebrain-specific alpha1(-/H) and alpha1(-/R) mice, respectively) in forebrain glutamatergic neurons. In the rest of the brain, alpha1(-/R) mutants are heterozygous alpha1(H101R) mice. Forebrain-specific alpha1(-/-) mice showed enhanced diazepam-induced motor depression and increased expression of the alpha2 and alpha3 subunits in the neocortex and hippocampus, in comparison with their pseudo-wild-type littermates. Forebrain-specific alpha1(-/R) mice were less sensitive than alpha1(-/H) mice to the motor-depressing action of diazepam, but each of these conditional mutants had a similar behavioral response as their corresponding control littermates. Unexpectedly, expression of the alpha1 subunit was reduced in forebrain, notably in alpha1(-/R) mice, and the alpha3 subunit was up-regulated in neocortex, indicating that proper alpha1 subunit expression requires both alleles. In conclusion, conditional manipulation of GABA(A) receptor alpha1 subunit expression can induce compensatory changes in the affected areas. Specifically, alterations in GABA(A) receptor expression restricted to forebrain glutamatergic neurons reproduce the behavioral effects seen after a global alteration, thereby implicating these neurons in the motor-sedative effect of diazepam.
Abstract Converging evidence from pharmacological and molecular studies has led to the suggestion that inhibition of glycine transporter 1 (GlyT1) constitutes an effective means to boost N -methyl- d ...-aspartate receptor (NMDAR) activity by increasing the extra-cellular concentration of glycine in the vicinity of glutamatergic synapses. However, the precise extent and limitation of this approach to alter cognitive function, and therefore its potential as a treatment strategy against psychiatric conditions marked by cognitive impairments, remain to be fully examined. Here, we generated mutant mice lacking GlyT1 in the entire forebrain including neurons and glia. This conditional knockout system allows a more precise examination of GlyT1 downregulation in the brain on behavior and cognition. The mutation was highly effective in attenuating the motor-stimulating effect of acute NMDAR blockade by phencyclidine, although no appreciable elevation in NMDAR-mediated excitatory postsynaptic currents (EPSC) was observed in the hippocampus. Enhanced cognitive performance was observed in spatial working memory and object recognition memory while spatial reference memory and associative learning remained unaltered. These findings provide further credence for the potential cognitive enhancing effects of brain GlyT1 inhibition. At the same time, they indicated potential phenotypic differences when compared with other constitutive and conditional GlyT1 knockout lines, and highlighted the possibility of a functional divergence between the neuronal and glia subpopulations of GlyT1 in the regulation of learning and memory processes. The relevance of this distinction to the design of future GlyT1 blockers as therapeutic tools in the treatment of cognitive disorders remains to be further investigated.
Neural networks in the spinal cord known as central pattern generators produce the sequential activation of muscles needed for locomotion. The overall locomotor network architectures in limbed ...vertebrates have been much debated, and no consensus exists as to how they are structured. Here, we use optogenetics to dissect the excitatory and inhibitory neuronal populations and probe the organization of the mammalian central pattern generator. We find that locomotor-like rhythmic bursting can be induced unilaterally or independently in flexor or extensor networks. Furthermore, we show that individual flexor motor neuron pools can be recruited into bursting without any activity in other nearby flexor motor neuron pools. Our experiments differentiate among several proposed models for rhythm generation in the vertebrates and show that the basic structure underlying the locomotor network has a distributed organization with many intrinsically rhythmogenic modules.
EphA4 signaling is essential for the spatiotemporal organization of neuronal circuit formation. In mice, deletion of this signaling pathway causes aberrant midline crossing of axons from both brain ...and spinal neurons and the complete knock-outs (KOs) exhibit a pronounced change in motor behavior, where alternating gaits are replaced by a rabbit-like hopping gait. The neuronal mechanism that is responsible for the gait switch in these KO mice is not known. Here, using intersectional genetics, we demonstrate that a spinal cord-specific deletion of EphA4 signaling is sufficient to generate the overground hopping gait. In contrast, selective deletion of EphA4 signaling in forebrain neurons, including the corticospinal tract neurons, did not result in a change in locomotor pattern. The gait switch was attributed to the loss of EphA4 signaling in excitatory Vglut2+ neurons, which is accompanied by an increased midline crossing of Vglut2+ neurons in the ventral spinal cord. Our findings functionally define spinal EphA4 signaling in excitatory Vglut2+ neurons as required for proper organization of the spinal locomotor circuitry, and place these cells as essential components of the mammalian locomotor network.
γ‐Aminobutyric acid (GABA)ergic neurons in the neocortex have been mainly regarded as interneurons and thought to provide local interactions. Recently, however, glutamate decarboxylase (GAD) ...immunocytochemistry combined with retrograde labeling experiments revealed the existence of GABAergic projection neurons in the neocortex. We further studied the network of GABAergic projection neurons in the neocortex by using GAD67‐green fluorescent protein (GFP) knock‐in mice for retrograde labeling and a novel neocortical GABAergic neuron labeling method for axon tracing. Many GFP‐positive neurons were retrogradely labeled after Fast Blue injection into the primary somatosensory, motor and visual cortices. These neurons were labeled not only around the injection site, but also at a long distance from the injection site. Of the retrogradely labeled GABAergic neurons remote from the injection sites, the vast majority (91%) exhibited somatostatin immunoreactivity, and were preferentially distributed in layer II, layer VI and in the white matter. In addition, most of GABAergic projection neurons were positive for neuropeptide Y (82%) and neuronal nitric oxide synthase (71%). We confirmed the long‐range projections by tracing GFP‐labeled GABAergic neurons with axon branches traveled rostro‐caudally and medio‐laterally. Axon branches could be traced up to 2 mm. Some (n = 2 of 4) were shown to cross the areal boundaries. The GABAergic projection neurons preferentially received neocortical inputs. From these results, we conclude that GABAergic projection neurons are distributed throughout the neocortex and are part of a corticocortical network.
The ephrin/Eph system plays a central role in neuronal circuit formation; however, its downstream effectors are poorly understood. Here we show that α-chimerin Rac GTPase-activating protein mediates ...ephrinB3/EphA4 forward signaling. We discovered a spontaneous mouse mutation,
miffy (
mfy), which results in a rabbit-like hopping gait, impaired corticospinal axon guidance, and abnormal spinal central pattern generators. Using positional cloning, transgene rescue, and gene targeting, we demonstrated that loss of α-chimerin leads to
mfy phenotypes similar to those of
EphA4
−/− and
ephrinB3
−/− mice. α-chimerin interacts with EphA4 and, in response to ephrinB3/EphA4 signaling, inactivates Rac, which is a positive regulator of process outgrowth. Moreover, downregulation of α-chimerin suppresses ephrinB3-induced growth cone collapse in cultured neurons. Our findings indicate that ephrinB3/EphA4 signaling prevents growth cone extension in motor circuit formation via α-chimerin-induced inactivation of Rac. They also highlight the role of a Rho family GTPase-activating protein as a key mediator of ephrin/Eph signaling.
Expanded polyglutamine (polyQ) proteins in Huntington’s disease (HD) as well as other polyQ disorders are known to elicit a variety of intracellular toxicities, but it remains unclear whether polyQ ...proteins can elicit pathological cell-cell interactions which are critical to disease pathogenesis. To test this possibility, we have created conditional HD mice expressing a neuropathogenic form of mutant huntingtin (mhtt-exon1) in discrete neuronal populations. We show that mhtt aggregation is a cell-autonomous process. However, progressive motor deficits and cortical neuropathology are only observed when mhtt expression is in multiple neuronal types, including cortical interneurons, but not when mhtt expression is restricted to cortical pyramidal neurons. We further demonstrate an early deficit in cortical inhibition, suggesting that pathological interactions between interneurons and pyramidal neurons may contribute to the cortical manifestation of HD. Our study provides genetic evidence that pathological cell-cell interactions elicited by neuropathogenic forms of mhtt can critically contribute to cortical pathogenesis in a HD mouse model.
In the rodent primary somatosensory cortex, the configuration of whiskers
and sinus hairs on the snout and of receptor-dense zones on the paws is topographically
represented as discrete modules of ...layer IV granule cells (barrels) and thalamocortical
afferent terminals. The role of neural activity, particularly
activity mediated by NMDARs (N-methyl-d-aspartate receptors),
in patterning of the somatosensory cortex has been a subject of debate. We have generated mice in which deletion of the NMDAR1
(NR1) gene is restricted to excitatory cortical neurons, and here
we show that sensory periphery-related patterns develop normally in the brainstem
and thalamic somatosensory relay stations of these mice. In the somatosensory
cortex, thalamocortical afferents corresponding to large whiskers form patterns
and display critical period plasticity, but their patterning is not as distinct
as that seen in the cortex of normal mice. Other thalamocortical patterns
corresponding to sinus hairs and digits are mostly absent. The cellular aggregates
known as barrels and barrel boundaries do not develop even at sites where
thalamocortical afferents cluster. Our findings indicate that cortical NMDARs
are essential for the aggregation of layer IV cells into barrels and for development
of the full complement of thalamocortical patterns.
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