GABAA Alpha 2,3 Modulation Improves Select Phenotypes in a Mouse Model of Fragile X Syndrome.

Fragile X syndrome (FXS) is the most common cause of inherited intellectual disability. FXS is caused by functional loss of the Fragile X Protein (FXP), also known as Fragile X Mental Retardation Protein (FMRP). In humans and animal models, loss of FXP leads to sensory hypersensitivity, increased susceptibility to seizures and cortical hyperactivity. Several components of the GABAergic system, the major inhibitory system in the brain, are dysregulated in FXS, and thus modulation of GABAergic transmission was suggested and tested as a treatment strategy. However, so far, clinical trials using broad spectrum GABAA or GABAB receptor-specific agonists have not yielded broad improvement of FXS phenotypes in humans. Here, we tested a more selective strategy in Fmr1 knockout (KO) mice using the experimental drug BAER-101, which is a selective GABAA α2/α3 agonist. Our results suggest that BAER-101 reduces hyperexcitability of cortical circuits, partially corrects increased frequency-specific baseline cortical EEG power, reduces susceptibility to audiogenic seizures and improves novel object memory. Other Fmr1 KO-specific phenotypes were not improved by the drug, such as increased hippocampal dendritic spine density, open field activity and marble burying. Overall, this work shows that BAER-101 improves select phenotypes in Fmr1 KO mice and encourages further studies into the efficacy of GABAA-receptor subunit-selective agonists for the treatment of FXS.

Acamprosate in a mouse model of fragile X syndrome: modulation of spontaneous cortical activity, ERK1/2 activation, locomotor behavior, and anxiety.

BACKGROUND: Fragile X Syndrome (FXS) occurs as a result of a silenced fragile X mental retardation 1 gene (FMR1) and subsequent loss of fragile X mental retardation protein (FMRP) expression. Loss of FMRP alters excitatory/inhibitory signaling balance, leading to increased neuronal hyperexcitability and altered behavior. Acamprosate (the calcium salt of N-acetylhomotaurinate), a drug FDA-approved for relapse prevention in the treatment of alcohol dependence in adults, is a novel agent with multiple mechanisms that may be beneficial for people with FXS. There are questions regarding the neuroactive effects of acamprosate and the significance of the molecule's calcium moiety. Therefore, the electrophysiological, cellular, molecular, and behavioral effects of acamprosate were assessed in the Fmr1-/y (knock out; KO) mouse model of FXS controlling for the calcium salt in several experiments. METHODS: Fmr1 KO mice and their wild-type (WT) littermates were utilized to assess acamprosate treatment on cortical UP state parameters, dendritic spine density, and seizure susceptibility. Brain extracellular-signal regulated kinase 1/2 (ERK1/2) activation was used to investigate this signaling molecule as a potential biomarker for treatment response. Additional adult mice were used to assess chronic acamprosate treatment and any potential effects of the calcium moiety using CaCl2 treatment on behavior and nuclear ERK1/2 activation. RESULTS: Acamprosate attenuated prolonged cortical UP state duration, decreased elevated ERK1/2 activation in brain tissue, and reduced nuclear ERK1/2 activation in the dentate gyrus in KO mice. Acamprosate treatment modified behavior in anxiety and locomotor tests in Fmr1 KO mice in which control-treated KO mice were shown to deviate from control-treated WT mice. Mice treated with CaCl2 were not different from saline-treated mice in the adult behavior battery or nuclear ERK1/2 activation. CONCLUSIONS: These data indicate that acamprosate, and not calcium, improves function reminiscent of reduced anxiety-like behavior and hyperactivity in Fmr1 KO mice and that acamprosate attenuates select electrophysiological and molecular dysregulation that may play a role in the pathophysiology of FXS. Differences between control-treated KO and WT mice were not evident in a recognition memory test or in examination of acoustic startle response/prepulse inhibition which impeded conclusions from being made about the treatment effects of acamprosate in these instances.

Comparison of Neurological Function in Males and Females from Two Substrains of C57BL/6 Mice.

The C57BL/6 (B6) mouse is the background strain most frequently used for genetically-modified mice. Previous studies have found significant behavioral and genetic differences between the B6J (The Jackson Laboratory) and B6N substrains (National Institutes of Health); however, most studies employed only male mice. We performed a comprehensive battery of motor function and learning and memory tests on male and female mice from both substrains. The B6N male mice had greater improvement in the rotarod test. In contrast, B6J female mice had longer latencies to falling from the rotarod. In the Morris water maze (MWM), B6J males had significantly shorter latencies to finding the hidden platform. However, B6N females had significantly shorter path lengths in the reversal and shifted-reduced phases. In open field locomotor activity, B6J males had higher activity levels, whereas B6N females took longer to habituate. In the fear conditioning test, B6N males had a significantly longer time freezing in the new context compared with B6J males, but no significant differences were found in contextual or cued tests. In summary, our findings demonstrate the importance of testing both males and females in neurobehavioral studies. Both factors (sex and substrain) must be taken into account when designing developmental neurotoxicology studies.

Ahrd Cyp1a2(-/-) mice show increased susceptibility to PCB-induced developmental neurotoxicity.

Polychlorinated biphenyls (PCBs) are developmental neurotoxicants that produce cognitive and behavioral changes in children exposed during gestation and lactation. Coplanar PCBs bind the aryl hydrocarbon receptor (AHR) and can be sequestered in liver by cytochrome P450 1A2 (CYP1A2). The AHR is a ligand-activated transcription factor which increases expression of the CYP1 family, including CYP1A2. Our previous work examining genetic susceptibility to developmental PCB neurotoxicity showed that Ahr(b)Cyp1a2(-/-) mice with the high-affinity Ahr(b) allele and lacking CYP1A2 were most susceptible while Ahr(b)Cyp1a2(+/+) and poor-affinity Ahr(d)Cyp1a2(+/+) mice were resistant. To follow up, a fourth line of mice was generated with the Ahr(d)Cyp1a2(-/-) genotype and compared with the background strain Ahr(b)Cyp1a2(+/+). Dams received a PCB mixture or the corn oil vehicle at gestational Day 10 (GD10) and postnatal Day 5 (PND5). Offspring were tested at PND60 in open field locomotor, acoustic startle with pre-pulse inhibition (PPI), novel object recognition and Morris water maze. Locomotor activity was increased in PCB-treated Ahr(b)Cyp1a2(+/+) mice, but no differences were seen in control vs. PCB-treated Ahr(d)Cyp1a2(-/-) mice. PCB-treated Ahr(d)Cyp1a2(-/-) mice had a higher baseline startle response and significantly reduced pre-pulse inhibition at the 74dB level compared with corn oil-treated controls (P<0.05). PCB-treated Ahr(d)Cyp1a2(-/-) mice had impairments in novel objective recognition (P<0.05) and during all three hidden platform phases of Morris water maze (P<0.01). Combined with our previous findings, these results indicate Cyp1a2 genotype is more important in susceptibility to PCB-induced deficits in learning and memory, but Ahr genotype appears more important when assessing acoustic startle-PPI and locomotor activity.