Cannabinoid CB 1 receptors (CB 1 Rs) mediate the presynaptic effects of endocannabinoids in the central nervous system (CNS) and most behavioral effects of exogenous cannabinoids. Cannabinoid receptor-interacting protein 1a (CRIP 1a ) binds to the CB 1 R C-terminus and can attenuate constitutive CB 1 R-mediated inhibition of Ca 21 channel activity. We now demonstrate cellular colocalization of CRIP 1a at neuronal elements in the CNS and show that CRIP 1a inhibits both constitutive and agonist-stimulated CB 1 Rmediated guanine nucleotide-binding regulatory protein (G-protein) activity. Stable overexpression of CRIP 1a in human embryonic kidney (HEK)-293 cells stably expressing CB 1 Rs (CB 1 -HEK), or in N18TG2 cells endogenously expressing CB 1 Rs, decreased CB 1 Rmediated G-protein activation (measured by agonist-stimulated [ activation. These effects were not attributable to differences in CB 1 R expression or endocannabinoid tone because CB 1 R levels did not differ between cell lines varying in CRIP 1a expression, and endocannabinoid levels were undetectable (CB 1 -HEK) or unchanged (N18TG2) by CRIP 1a overexpression. In CB 1 -HEK cells, 4-hour pretreatment with cannabinoid agonists downregulated CB 1 Rs and desensitized agonist-stimulated [ 35 S]GTPgS binding. CRIP 1a overexpression attenuated CB 1 R downregulation without altering CB 1 R desensitization. Finally, in cultured autaptic hippocampal neurons, CRIP 1a overexpression attenuated both depolarizationinduced suppression of excitation and inhibition of excitatory synaptic activity induced by exogenous application of cannabinoid but not by adenosine A1 agonists. These results confirm that CRIP 1a inhibits constitutive CB 1 R activity and demonstrate that CRIP 1a can also inhibit agonist-stimulated CB 1 R signaling and downregulation of CB 1 Rs. Thus, CRIP 1a appears to act as a broad negative regulator of CB 1 R function.

Summary Purpose The endocannabinoid system is known to modulate seizure activity in several in vivo and in vitro models, and CB1-receptor activation is anticonvulsant in the rat pilocarpine model of acquired epilepsy (AE). In these epileptic rats, a unique redistribution of the CB1 receptor occurs within the hippocampus; however, an anatomically inclusive analysis of the effect of status epilepticus (SE)–induced AE on CB1 receptors has not been thoroughly evaluated. Therefore, statistical parametric mapping (SPM), a whole-brain unbiased approach, was used to study the long-term effect of pilocarpine-induced SE on CB1-receptor binding and G-protein activation in rats with AE. Methods Serial coronal sections from control and epileptic rats were cut at equal intervals throughout the neuraxis and processed for [3H]WIN55,212-2 (WIN) autoradiography, WIN-stimulated [35S]GTPγS autoradiography, and CB1-receptor immunohistochemistry (IHC). The autoradiographic techniques were evaluated with both region of interest (ROI) and SPM analyses. Key Findings In rats with AE, regionally specific increases in CB1-receptor binding and activity were detected in cortex, discrete thalamic nuclei, and other regions including caudate-putamen and septum, and confirmed by IHC. However, CB1 receptors were unaltered in several brain regions, including substantia nigra and cerebellum, and did not exhibit regional decreases in rats with AE. Significance This study provides the first comprehensive evaluation of the regional distribution of changes in CB1-receptor expression, binding, and G-protein activation in the rat pilocarpine model of AE. These regions may ultimately serve as targets for cannabinomimetic compounds or manipulation of the endocannabinoid system in epileptic brain.