Addressing sufficiency of the CB1 receptor for endocannabinoid-mediated functions through conditional genetic rescue in forebrain GABAergic neurons

Abstract

Genetic inactivation of the cannabinoid CB1 receptor gene in different cell types in the brain has previously revealed necessary functions for distinct synaptic plasticity processes and behaviors. Here, we sought to identify CB1 receptor expression sites that are minimally required to reconstruct normal phenotypes. In a CB1-null background, we re-expressed endogenous CB1 receptors in forebrain GABAergic neurons, thereby assessing the sufficiency of CB1 receptors. Depolarization-induced suppression of inhibitory, but not excitatory, transmission was restored in hippocampal and amygdalar circuits. GABAergic CB1 receptors did not convey protection against chemically induced seizures, but prevented the spontaneous mortality observed in CB1 null mutants. Rescue of GABAergic CB1 receptors largely restored normal anxiety-like behavior but improved extinction of learned fear only marginally. This study illustrates that the approach of genetic reconstruction of complex behaviors is feasible. It also revealed distinct degrees of modulation for different emotional behaviors by the GABAergic population of CB1 receptors.Electronic supplementary materialThe online version of this article (doi:10.1007/s00429-017-1411-5) contains supplementary material, which is available to authorized users.

Localization of the cannabinoid type‐1 receptor in subcellular astrocyte compartments of mutant mouse hippocampus

“…Specificity control experiments of the CB 1 receptor antibodies were carried out in CB 1 ‐KO and STOP‐ CB 1 mice (carrying a loxP‐flanked stop cassette inserted into the sequences of the 5'UTR of the CB 1 receptor). According to recent observations, we detected very low levels of metal particle deposits in STOP‐ CB 1 (Remmers et al, ) and scattered background particles in CB 1 ‐KO.…”

“…The GFAP‐ CB 1 ‐RS mouse expressing CB 1 receptors exclusively in astrocytes described here, together with the Glu‐ CB 1 ‐RS rescue mouse expressing the receptor only in dorsal telencephalic glutamatergic neurons (de Salas‐Quiroga et al, ; Lange et al, ; Ruehle et al, ; Soria‐Gómez et al, ) and the GABA‐ CB 1 ‐RS rescue mouse expressing the CB 1 receptor only in GABAergic neurons (de Salas‐Quiroga et al, ; Lange et al, ) that were recently characterized anatomically (Gutiérrez‐Rodríguez et al, ; Remmers et al, ), suggest that the regulation of the CB 1 receptor expression in astrocytes, glutamatergic neurons and GABAergic neurons may be independent of each others. The present demonstration that the GFAP‐ CB 1 ‐RS in the hippocampus maintains the normal CB 1 receptor expression and distribution in astrocytes make these mutants ideal suited for the study of the astroglial CB 1 receptor function, as shown for Glu‐ CB 1 ‐RS (de Salas‐Quiroga et al, ; Gutiérrez‐Rodríguez et al, ; Lange et al, ; Ruehle et al, ; Soria‐Gómez et al, ) and GABA‐ CB 1 ‐RS mice (de Salas‐Quiroga et al, ; Gutiérrez‐Rodríguez et al, ; Lange et al, ; Remmers et al, ). In fact, these rescue strategies have the advantage of the restoration and visualization of existing CB 1 receptor levels in locations with sparse CB 1 receptors (as the astrocytes and astroglial mitochondria), allowing a more comprehensive functional characterization of the (endo)cannabinoid system based on the precise cellular and subcellular localization of the CB 1 receptor.…”

“…The rescue of CB 1 receptors in mice expressing the gene exclusively in dorsal telencephalic glutamatergic neurons (Glu‐ CB 1 ‐RS) or in forebrain GABAergic neurons (GABA‐ CB 1 ‐RS) (de Salas‐Quiroga et al, ; Lange et al, ; Remmers et al, ; Ruehle et al, ; Soria‐Gómez et al, ) has provided interesting insights into the sufficiency of the CB 1 receptor in these cells for specific brain functions and behaviors. Therefore, restoration of CB 1 receptor expression in astrocytes and astroglial mitochondria could represent a new approach to assess the function of the tripartite synapse.…”

“…These effects are due to intra‐mitochondrial Gαi protein activation by mtCB 1 receptors that leads to the inhibition of soluble adenylyl cyclase and, consequently, to the decrease in intra‐mitochondrial protein kinase A (PKA) activity (Hebert‐Chatelain et al, ). New tools based on genetic rescue strategies have proven to be useful to dissect the sufficiency of the CB 1 receptors expressed in specific cell types for a particular brain function (de Salas‐Quiroga et al, ; Gutiérrez‐Rodríguez et al, ; Lange et al, ; Remmers et al, ; Ruehle et al, ; Soria‐Gómez et al, ). Importantly, knock‐in mice with cell type‐specific rescue of CB 1 receptors in dorsal telencephalic glutamatergic neurons (Glu‐ CB 1 ‐RS) or GABAergic neurons (GABA‐ CB 1 ‐RS) showed that the distribution pattern and the subcellular CB 1 receptor localization is maintained as it is observed in the wild‐type hippocampus (Gutiérrez‐Rodríguez et al, ; Remmers et al, ).…”

Cannabinoid receptor 1/2 double‐knockout mice develop epilepsy

“…Finally, although Cnr1 −/− mice had similar mortality to double‐knockouts, they did not show spontaneous seizure activity, suggesting that early mortality is driven by factors other than seizures. Other groups have reported increased mortality in Cnr1 −/− mice, but the cause remains unclear. Cardiovascular or other neurological problems besides seizures could lead to the observed increase in mortality.…”