Aberrant Somatosensory Processing and Connectivity in Mice Lacking Engrailed-2.

Overreactivity and defensive behaviors in response to tactile stimuli are common symptoms in autism spectrum disorder (ASD) patients. Similarly, somatosensory hypersensitivity has also been described in mice lacking ASD-associated genes such as Fmr1 (fragile X mental retardation protein 1). Fmr1 knock-out mice also show reduced functional connectivity between sensory cortical areas, which may represent an endogenous biomarker for their hypersensitivity. Here, we measured whole-brain functional connectivity in Engrailed-2 knock-out (En2-/-) adult mice, which show a lower expression of Fmr1 and anatomical defects common to Fmr1 knock-outs. MRI-based resting-state functional connectivity in adult En2-/- mice revealed significantly reduced synchronization in somatosensory-auditory/associative cortices and dorsal thalamus, suggesting the presence of aberrant somatosensory processing in these mutants. Accordingly, when tested in the whisker nuisance test, En2-/- but not WT mice of both sexes showed fear behavior in response to repeated whisker stimulation. En2-/- mice undergoing this test exhibited decreased c-Fos-positive neurons (a marker of neuronal activity) in layer IV of the primary somatosensory cortex and increased immunoreactive cells in the basolateral amygdala compared with WT littermates. Conversely, when tested in a sensory maze, En2-/- and WT mice spent a comparable time in whisker-guided exploration, indicating that whisker-mediated behaviors are otherwise preserved in En2 mutants. Therefore, fearful responses to somatosensory stimuli in En2-/- mice are accompanied by reduced basal connectivity of sensory regions, reduced activation of somatosensory cortex, and increased activation of the basolateral amygdala, suggesting that impaired somatosensory processing is a common feature in mice lacking ASD-related genes.SIGNIFICANCE STATEMENT Overreactivity to tactile stimuli is a common symptom in autism spectrum disorder (ASD) patients. Recent studies performed in mice bearing ASD-related mutations confirmed these findings. Here, we evaluated the behavioral response to whisker stimulation in mice lacking the ASD-related gene Engrailed-2 (En2-/- mice). Compared with WT controls, En2-/- mice showed reduced functional connectivity in the somatosensory cortex, which was paralleled by fear behavior, reduced activation of somatosensory cortex, and increased activation of the basolateral amygdala in response to repeated whisker stimulation. These results suggest that impaired somatosensory signal processing is a common feature in mice harboring ASD-related mutations.

Epileptiform activity and cognitive deficits in SNAP-25(+/-) mice are normalized by antiepileptic drugs.

Synaptosomal-associated protein of 25 kDa (SNAP-25) is a protein that participates in the regulation of synaptic vesicle exocytosis through the formation of the soluble NSF attachment protein receptor complex and modulates voltage-gated calcium channels activity. The Snap25 gene has been associated with schizophrenia, attention deficit hyperactivity disorder, and bipolar disorder, and lower levels of SNAP-25 have been described in patients with schizophrenia. We used SNAP-25 heterozygous (SNAP-25(+/-)) mice to investigate at which extent the reduction of the protein levels affects neuronal network function and mouse behavior. As interactions of genotype with the specific laboratory conditions may impact behavioral results, the study was performed through a multilaboratory study in which behavioral tests were replicated in at least 2 of 3 distinct European laboratories. Reductions of SNAP-25 levels were associated with a moderate hyperactivity, which disappeared in the adult animals, and with impaired associative learning and memory. Electroencephalographic recordings revealed the occurrence of frequent spikes, suggesting a diffuse network hyperexcitability. Consistently, SNAP-25(+/-) mice displayed higher susceptibility to kainate-induced seizures, paralleled by degeneration of hilar neurons. Notably, both EEG profile and cognitive defects were improved by antiepileptic drugs. These results indicate that reduction of SNAP-25 expression is associated to generation of epileptiform discharges and cognitive dysfunctions, which can be effectively treated by antiepileptic drugs.

Antiepileptic effects of botulinum neurotoxin E.

Experimental studies suggest that the delivery of antiepileptic agents into the seizure focus might be of potential utility for the treatment of focal-onset epilepsies. Botulinum neurotoxin E (BoNT/E) causes a prolonged inhibition of neurotransmitter release after its specific cleavage of the synaptic protein synaptosomal-associated protein of 25 kDa (SNAP-25). Here, we show that BoNT/E injected into the rat hippocampus inhibits glutamate release and blocks spike activity of pyramidal neurons. BoNT/E effects persist for at least 3 weeks, as determined by immunodetection of cleaved SNAP-25 and loss of intact SNAP-25. The delivery of BoNT/E to the rat hippocampus dramatically reduces both focal and generalized kainic acid-induced seizures as documented by behavioral and electrographic analysis. BoNT/E treatment also prevents neuronal loss and long-term cognitive deficits associated with kainic acid seizures. Moreover, BoNT/E-injected rats require 50% more electrical stimulations to reach stage 5 of kindling, thus indicating a delayed epileptogenesis. We conclude that BoNT/E delivery to the hippocampus is both antiictal and antiepileptogenic in experimental models of epilepsy.

Simultaneous absence of dopamine D1 and D2 receptor-mediated signaling is lethal in mice.

Dopamine (DA) controls a wide variety of physiological functions in the central nervous system as well as in the neuroendocrine and gastrointestinal systems. DA signaling is mediated by five cloned receptors named D1-D5. Knockout mouse models for the five receptors have been generated, and, albeit impaired for some important DA-mediated functions, they are viable and can reproduce. D1 and D2 receptors are the most abundant and widely expressed DA receptors. Cooperative/synergistic effects mediated by these receptors have been suggested, in particular, in the control of motor behaviors. To analyze the extent of such interrelationship, we have generated double D1/D2 receptor mutants. Interestingly, in contrast to single knockouts, we found that concurrent ablation of the D1 and D2 receptors is lethal during the second or third week after birth. This dramatic phenotype is likely to be related to altered feeding behavior and dysfunction of the gastrointestinal system, especially because major anatomical changes were not identified in the brain. Similarly, in the absence of functional D1, heterozygous D2 mutants (D1r(-/-);D2r(+/-)) showed severe growth retardation and did not survive their postweaning period. The analysis of motor behavior in D1r/D2r compound mutants showed that loss of D2-mediated functions reduces motor abilities, whereas the effect of D1r ablation on locomotion strongly depends on the experimental paradigms used. These studies highlight the interrelationship between D1 and D2 receptor-mediated control of motor activity, food intake, and gastrointestinal functions, which has been elusive in the single-gene ablation studies.