Localization of cannabinoid CB1 receptor mRNA in neuronal subpopulations of rat striatum: A double-label in situ hybridization study

Double-label in situ hybridization was used to identify the phenotypes of s triatal neurons that express mRNA for cannabinoid CB, receptors. Simultaneo us detection of multiple mRNAs was performed by combining a S-35-labeled ri bonucleotide probe for CB, mRNA with digoxigenin-labeled riboprobes for str iatal projection neurons (preprotachykinin A, prodynorphin, and preproenkep halin mRNAs) and interneurons (vesicular acetylcholine transporter (VAChT), choline acetyltransferase (ChAT), somatostatin, and glutamic acid decarbox ylase (Mr 67,000; GAD67) mRNAs). To ascertain whether CB1 mRNA was a marker for striatal efferents, digoxigenin-labeled probes for mRNA markers of bot h striatonigral (prodynorphin or preprotachykinin A mRNAs), and striatopall idal (proenkephalin mRNAs) projection neurons were combined with the 35S-la beled probe for CB1. A mediolateral gradient in CB1 mRNA expression was obs erved at rostral and mid-striatal levels; in the same coronal sections the number of silver grains per cell ranged from below the threshold of detecta bility at the medial and ventral poles to saturation at the dorsolateral bo undary bordered by the corpus callosum. At the caudal level examined, CB1 m RNA was denser in the ventral sector relative to the dorsal sector. Virtual ly all neurons expressing mRNA markers for striatal projection neurons colo calized CB1 mRNA. Combining a S-35-labeled riboprobe for CB1 with digoxigen in-labeled riboprobes for both preproenkephalin and prodynorphin confirmed localization of CB1 mRNA to striatonigral and striatopallidal neurons expre ssing prodynorphin and preproenkephalin mRNAs, respectively. However, CB1 m RNA-positive cells that failed to coexpress the other markers were also app arent. CB1 mRNA was localized to putative GABAergic interneurons that expre ss high levels of GAD67 mRNA. These interneurons enable functional interact ions between the direct and indirect striatal output pathways. By contrast, aspiny interneurons that express preprosomatostatin mRNA and cholinergic i nterneurons that coexpress ChAT and VAChT mRNAs were CB1 mRNA-negative, The present data provide direct evidence that cannabinoid receptors are synthe sized in striatonigral neurons that contain dynorphin and substance P and s triatopallidal neurons that contain enkephalin. By contrast, local circuit neurons in striatum that contain somatostatin or acetylcholine do not synth esize cannabinoid receptors. Published 2000 Wiley-Liss, Inc(dagger).