Fatty acid amide hydrolase (FAAH) and monoglyceride lipase (MGL) catalyse the hydrolysis of the endocannabinoids anandamide and 2-arachidonoyl glycerol. We investigated their ultrastructural distribution in brain areas where the localization and effects of cannabinoid receptor activation are known. In the hippocampus, FAAH was present in somata and dendrites of principal cells, but not in interneurons. It was located mostly on the membrane surface of intracellular organelles known to store Ca 2+ (e.g. mitochondria, smooth endoplasmic reticulum), less frequently on the somatic or dendritic plasma membrane. MGL immunoreactivity was found in axon terminals of granule cells, CA3 pyramidal cells and some interneurons. In the cerebellum, Purkinje cells and their dendrites are intensively immunoreactive for FAAH, together with a sparse axon plexus at the border of the Purkinje cell ⁄ granule cell layers. Immunostaining for MGL was complementary, the axons in the molecular layer were intensively labelled leaving the Purkinje cell dendrites blank. FAAH distribution in the amygdala was similar to that of the CB 1 cannabinoid receptor: evident signal in neuronal somata and proximal dendrites in the basolateral nucleus, and hardly any labelling in the central nucleus. MGL staining was restricted to axons in the neuropil, with similar relative signal intensities seen for FAAH in different nuclei. Thus, FAAH is primarily a postsynaptic enzyme, whereas MGL is presynaptic. FAAH is associated with membranes of cytoplasmic organelles. The differential compartmentalization of the two enzymes suggests that anandamide and 2-AG signalling may subserve functional roles that are spatially segregated at least at the stage of metabolism.

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Naϩ /Ca 2ϩ exchanger 3 (NCX3), one of the three isoforms of the NCX family, is highly expressed in the brain and is involved in the maintenance of intracellular Na ϩ and Ca 2ϩ homeostasis. Interestingly, whereas the function of NCX3 under physiological conditions has been determined, its role under anoxia is still unknown. To assess NCX3 role in cerebral ischemia, we exposed ncx3Ϫ/Ϫ mice to transient middle cerebral artery occlusion followed by reperfusion. In addition, to evaluate the effect of ncx3 ablation on neuronal survival, organotypic hippocampal cultures and primary cortical neurons from ncx3Ϫ/Ϫ mice were subjected to oxygen glucose deprivation (OGD) plus reoxygenation. Here we report that ncx3 gene suppression leads to a worsening of brain damage after focal ischemia and to a massive neuronal death in all the hippocampal fields of organotypic cultures as well as in cortical neurons from ncx3Ϫ/Ϫ mice exposed to OGD plus reoxygenation. In addition, in ncx3Ϫ/Ϫ cortical neurons exposed to hypoxia, NCX currents, recorded in the reverse mode of operation, were significantly lower than those detected in ncx3ϩ/ϩ. From these results, NCX3 protein emerges as a new molecular target that may have a potential therapeutic value in modulating cerebral ischemia.

In the central nervous system (CNS), the Na(+)-Ca(2+) exchanger plays a fundamental role in controlling the changes in the intracellular concentrations of Na(+) and Ca(2+) ions. These cations are known to regulate neurotransmitter release, cell migration and differentiation, gene expression, and neurodegenerative processes. In the present study, nonradioactive in situ hybridization and light immunohistochemistry were carried out to map the regional and cellular distribution for both transcripts and proteins encoded by the three known Na(+)-Ca(2+) exchanger genes NCX1, NCX2, and NCX3. NCX1 transcripts were particularly expressed in layers III-V of the motor cortex, in the thalamus, in CA3 and the dentate gyrus of the hippocampus, in several hypothalamic nuclei, and in the cerebellum. NCX2 transcripts were strongly expressed in all hippocampal subregions, in the striatum, and in the paraventricular thalamic nucleus. NCX3 mRNAs were mainly detected in the hippocampus, in the thalamus, in the amygdala, and in the cerebellum. Immunohistochemical analysis revealed that NCX1 protein was mainly expressed in the supragranular layers of the cerebral cortex, in the hippocampus, in the hypothalamus, in the substantia nigra and ventral tegmental area, and in the granular layer of the cerebellum. The NCX2 protein was predominantly expressed in the hippocampus, in the striatum, in the thalamus, and in the hypothalamus. The NCX3 protein was particularly found in the CA3 subregion, and in the oriens, radiatum, and lacunoso-moleculare layers of the hippocampus, in the ventral striatum, and in the cerebellar molecular layer. Collectively, these results suggest that the different Na(+)-Ca(2+) exchanger isoforms appear to be selectively expressed in several CNS regions where they might underlie different functional roles.

Voltage-dependent K(+) channels play a pivotal role in controlling cellular excitability within the nervous system. The aim of the present study was to investigate the expression in the adult rat brain of the three ether-a-gogo-related gene (ERG) family members ERG1, ERG2, and ERG3, encoding for K(+) channel subunits. To this aim, the distribution of ERG transcripts was studied by means of reverse-transcription polymerase chain reaction (RT-PCR) and nonradioactive in situ hybridization histochemistry (NR-ISH). Furthermore, ERG1 subunit distribution was studied by immunohistochemical analysis. RT-PCR analysis revealed ERG1, ERG2, and ERG3 expression in the olfactory bulb, cerebral cortex, hippocampus, hypothalamus, and cerebellum. NR-ISH experiments detected transcripts encoded by all three ERG genes in the cerebral cortex and in all CA subfields and in the granular cell layer of the dentate gyrus of the hippocampus; strong ERG1 signals were also detected in scattered large elements throughout the oriens, pyramidal, and radiatum layers, and in the hilus of the dentate gyrus. In the thalamus, positively labeled neurons were detected in the reticular nucleus with ERG1 and ERG3 and in the anterodorsal nucleus with ERG2 riboprobes. Transcripts for ERG1 and, to a lesser degree, also for ERG3, were detected in the basal ganglia and in several brainstem nuclei. All three ERG genes appeared to be expressed in cerebellar Purkinje cells. Finally, ERG1 expression was also revealed in non-neuronal elements such as ependymal and subependymal cells along the ventricular walls and hippocampal astrocytes. These results suggest that the K(+) channel isoforms of the ERG family appear to be expressed in different central nervous system regions where they might differentially control the firing of neurons engaged in several networks.