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  • Addiction, Drugs
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    719—Genetic Models of Autism in Animals

    Wednesday, November 13, 2013, 8:00 am - 12:00 noon

    719.03: Electrophysiological characterization of the Cntnap2/CASPR2 knockout mouse, a model for ASD

    Location: Halls B-H

    1Neurosci., 2Program in Neurogenetics, Dept. of Neurology, David Geffen Sch. of Med., 3Dept. of Neurology, David Geffen Sch. of Med., Univ. of California, Los Angeles, CA

    Abstract Body: Recent large-scale gene sequencing studies in autism spectrum disorder (ASD) demonstrate that a large number of genes are potentially involved in disease etiology. Recessive mutations in one of these genes, contactin associated protein-like 2 (CNTNAP2; called CASPR2 in mice), cause cortical dysplasia focal epilepsy syndrome (CDFE), a syndromic form of ASD in humans. Previously published characterization of the Cntnap2/CASPR2 knock-out (KO) mouse revealed striking parallels with the human behavioral phenotype (Peñagarikano et al. 2011, Cell), including the triad of repetitive/restrictive behavior, impaired language and social interactions. Seizures, decreased neuronal synchronization, decreased interneurons and neuronal migration deficits were also observed. To obtain a better understanding of the mechanisms underlying these phenotypes, we performed whole-cell recordings from L2/3 excitatory (WT n=31, KO n=25) and parvalbumin (PV+) interneurons (WT n=17, KO n=19) in in-vitro slices from the prelimbic medial prefrontal cortex. Intrinsic excitability, assessed by measuring the number of spikes elicited by increasing current steps was normal in excitatory neurons, but was diminished in PV+ neurons in Cntnap2 KO mice compared to controls. Moreover, voltage-clamp recordings of inhibitory and excitatory miniature postsynaptic currents (mIPSCs and mEPSC ) in pyramidal cells revealed a two-fold decrease in the frequency of mEPSCs, but no changes in mEPSC amplitude (WT n=19, KO n=22). We also observed a 30% decrease in mIPSC frequency (WT n=23, KO n=22). Therefore, deletion of Cntnap2 results in a dramatic decrease in functional excitatory inputs and a smaller decrease in inhibitory inputs, changing the balance of excitation and inhibition. This likely arises form decreased dendritic arbors and spine growth, and is consistent with a previous in vitro study (Anderson et al. 2012, PNAS). The changes in intrinsic excitability on the other hand suggest that CNTNAP2’s function in the axon initial segment may also play a role in this pathology, although the reason why this effect seems specific to PV+ interneurons remains unknown. Future studies will be focused on characterizing the mechanisms that result in these changes. Further work is needed in order to elucidate how physiological alterations in the model lead to pathological changes in network activity, as well as to gauge the effect of targeted therapeutic approaches on brain physiology and behavior.

    Lay Language Summary: We have studied a mouse that has been genetically engineered with reduced CASPR2/Cntnap2, similar to patients with mutations in this gene that causes autism. We discovered that neurons in the brain of these mice made fewer connections with each other. Moreover, we found that a specific group neurons, called parvalbumin interneurons, which are necessary for rapidly inhibiting the activity of other neurons and therefore keeping brain activity in balance, are less excitable, as they fire less action potentials in response to the same input.
    Individuals diagnosed with Autism Spectrum Disorder (ASD) show deficits in language and social interactions, as well as repetitive or restrictive behaviors. In some cases, ASD is accompanied by hyperactivity, attention deficits, and epileptic seizures. It is currently estimated that 1 in every 88 individuals has autism in the United States, a number that appears to be increasing and yet, there are none or few effective treatments for the disorder. Recent studies have shown that at least 20 percent of autism cases can be caused by genetic mutations in our DNA. One of these autism-causing genes, CNTNAP2, is involved in brain development and neuronal migration. We found that transgenic mice that lack this gene recapitulate the triad of behavioral deficits of ASD: decreased socialization, decreased vocalizations, and decreased grooming, as well as seizures and a reduction in inhibitory neurons.
    We then wanted to understand how alterations in the pattern of connections made by prefrontal neurons and changes in their excitability could lead to the disorder. This could give us clues about how a specific genetic cause can alter brain activity and lead to changes in behavior. Therefore, we recorded neuronal activity from neurons in brain slices of mice lacking Cntnap2, as well as in normal mice. These recordings were done specifically in the medial prefrontal cortex, an area of the brain associated with social behavior. This technique enabled us to quantify the amount of synaptic inputs or connections that each cell receives, as well to quantify their ability to send nerve impulses. We also recorded electrical activity from two neuronal subtypes: excitatory neurons and parvalbumin-positive inhibitory neurons.
    When compared to normal mice, we found that excitatory neurons in mice lacking Cntnap2 received less functional excitatory synaptic inputs. We also found that inhibitory interneurons fired fewer action potentials in response to similar stimuli. Interestingly, the number of excitatory inputs onto inhibitory neurons was unchanged. In addition, there was no change in the ability of excitatory neurons to fire action potentials and the number of excitatory inputs onto inhibitory neurons was unchanged.
    Taken as a whole, these findings support the idea that a change in connectivity between neurons in the prefrontal cortex could potentially lead to alterations in brain function and behavior, very much like those observed in autistic individuals.
    Our research shows that understanding how specific genes, like CNTNAP2, cause autism in humans, can be useful in revealing mechanisms that give rise to the disorder. The next step will be to determine how activity patterns of a population of interconnected neurons (brain network activity) is affected in these animals during a social behavioral task. This research will be crucial for establishing targeted treatments for ASD, as well as gauging the effect of therapeutic approaches.