Components of Endocannabinoid Signaling System are Expressed in the Perinatal Mouse Cerebellum and Required for its Normal Development.

Endocannabinoid signaling system (ECS), encompassing cannabinoid receptors and enzymes involved in the synthesis and degradation of the endogenous cannabinoid signaling lipids, is highly expressed in the cerebellar cortex of adult humans and rodents. In addition to their well-established role in neuromodulation, endocannabinoids (eCBs) have been shown to play key roles in aspects of neurodevelopment in the fore- and mid-brain, including neurogenesis, cell migration, and synapse specification. However, little is known about the role of ECS in cerebellar development. In this study we carried out immunohistochemical characterization of ECS components through key stages of cerebellar development in mice utilizing antibodies for 2-arachidonoylglycerol synthetizing and degrading enzymes and the major brain cannabinoid receptor, cannabinoid receptor 1 (CB1), in combination with cerebellar cell markers. Our results reveal a temporally, spatially, and cytologically dynamic pattern of expression. Production, receptor binding, and degradation of eCBs are tightly controlled, thus localization of eCB receptors and the complementary cannabinoid signaling machinery determines the direction, duration, and ultimately the outcome of eCB signaling. To gain insights into the role of eCB signaling in cerebellar development, we characterized gross anatomy of cerebellar midvermis in CB1 knockout (CB1 KO) mice, as well as their performance in cerebellar-influenced motor tasks. Our results show persistent and selective anatomical and behavioral alterations in CB1 KOs. Consequently, the insights gained from this study lay down the foundation for investigating specific cellular and molecular mechanisms regulated by eCB signaling during cerebellar development.

SIGNIFICANCE In this study we show that components of the endocannabinoid signaling system are prominently expressed in the perinatal (E17.5-P14) mouse hindbrain. Our comprehensive characterization highlights developmentally dynamic and spatially restricted expression of cannabinoid receptor 1, prominent at birth in pontocerebellar axons, and later in migrating and differentiating anterior vermis granule cells. We identify the role of Purkinje cells in the regulation of endocannabinoid 2-AG availability, since they express both catabolic and anabolic enzymes DAGLα and MAGL. Furthermore, we demonstrate a requirement for endocannabinoid signaling in the regulation of pontocerebellar axon distribution and of postnatal cerebellar growth. CB1 knockouts exhibit impairments in cerebellar-influenced fine-motor, but not gross-motor behaviors. Together, these results illuminate a previously unrecognized role of endocannabinoid signaling in the regulation of cerebellar development and function.

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