Abstract
The CB1 cannabinoid receptor is the most abundant G-protein coupled receptor in the brain and a key regulator of neuronal excitability. There is strong evidence that CB1 receptor on glutamatergic hippocampal neurons is beneficial to alleviate epileptiform seizures in mouse and man. Therefore, we hypothesized that experimentally increased CB1 gene dosage in principal neurons would have therapeutic effects in kainic acid (KA)-induced hippocampal pathogenesis. Here, we show that virus-mediated conditional overexpression of CB1 receptor in pyramidal and mossy cells of the hippocampus is neuroprotective and moderates convulsions in the acute KA seizure model in mice. We introduce a recombinant adeno-associated virus (AAV) genome with a short stop element flanked by loxP sites, for highly efficient attenuation of transgene expression on the transcriptional level. The presence of Cre-recombinase is strictly necessary for expression of reporter proteins or CB1 receptor in vitro and in vivo. Transgenic CB1 receptor immunoreactivity is targeted to glutamatergic neurons after stereotaxic delivery of AAV to the dorsal hippocampus of the driver mice NEX-cre. Increased CB1 receptor protein levels in hippocampal lysates of AAV-treated Cre-mice is paralleled by enhanced cannabinoid-induced G-protein activation. KA-induced seizure severity and mortality is reduced in CB1 receptor overexpressors compared with AAV-treated control animals. Neuronal damage in the hippocampal CA3 field is specifically absent from AAV-treated Cre-transgenics, but evident throughout cortical areas of both treatment groups. Our data provide further evidence for a role of increased CB1 signaling in pyramidal hippocampal neurons as a safeguard against the adverse effects of excessive excitatory network activity.
Highlights
Maintaining an optimal balance between excitatory and inhibitory activity of central nervous system (CNS) neurons is essential for proper physiological network activities, since either excessive glutamatergic transmission or insufficient GABAergic transmission can lead to excitotoxicity and epileptiform seizures in rodents and man [1]
The endocannabinoid system has been implied as a therapeutical target in epilepsy [11] and as such, effective treatment strategies utilizing cannabinoid type 1 (CB1) receptor regulation require a detailed understanding of CB1 receptor effects in neuronal subtypes
The analysis of conditional mouse mutants lacking CB1 receptors on different subtypes of neurons subjected to the kainic acid (KA)-induced seizures revealed that CB1 receptors on hippocampal glutamatergic but not GABAergic neurons are required for protection against excitotoxic seizures [10]
Summary
Maintaining an optimal balance between excitatory and inhibitory activity of central nervous system (CNS) neurons is essential for proper physiological network activities, since either excessive glutamatergic transmission or insufficient GABAergic transmission can lead to excitotoxicity and epileptiform seizures in rodents and man [1]. The endocannabinoid system has been implied as a therapeutical target in epilepsy [11] and as such, effective treatment strategies utilizing CB1 receptor regulation require a detailed understanding of CB1 receptor effects in neuronal subtypes To this end, the analysis of conditional mouse mutants lacking CB1 receptors on different subtypes of neurons subjected to the kainic acid (KA)-induced seizures revealed that CB1 receptors on hippocampal glutamatergic but not GABAergic neurons are required for protection against excitotoxic seizures [10]. The analysis of conditional mouse mutants lacking CB1 receptors on different subtypes of neurons subjected to the kainic acid (KA)-induced seizures revealed that CB1 receptors on hippocampal glutamatergic but not GABAergic neurons are required for protection against excitotoxic seizures [10] In line with this pre-clinical data, specific down-regulation of protein and mRNA of CB1 receptor on glutamatergic, but not on GABAergic axon terminals had been reported in epileptic human hippocampal tissue [12]. These conditional loss-offunction studies have not yet been complemented by the corresponding gain-of function approach entailing CB1 overexpression, preventing a comprehensive picture of CB1-mediated control of overexcitation
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