Cytoarchitectural relationships between [ 3H]ouabain binding and mRNA for isoforms of the sodium pump catalytic subunit in rat brain

Abstract

We examined the cell type-specific expression of the α1, α2, and α3 subunits of the sodium pump in rat brain using in situ hybridization and [ 3H]ouabain autoradiography. These techniques allowed us to colocalize mRNA and functional α2 α3 pumps on adjacent sections. The perikarya of many neurons possessed high levels of α1 and/or α3 transcripts, while α2 mRNA appeared to be present in only a few neuronal types. [ 3H]Ouabain binding in general paralleled the distribution of α3 mRNA-positive neurons. The regional variation of α1 and α3 transcripts was complex and varied. Large neurons of the olfactory bulb and piriform cortex expressed high levels of α3 transcripts, but low levels of α1 mRNA. In frontal cortex, neurons of layers II–III were enriched in α1 mRNA, while those in layer V exhibited high levels of α3 transcripts. In the hippocampus, principal neurons expressed all three α subunit mRNAs. CA subfield pyramidal neurons exhibited a high α3 α1 ratio, while dentate granule cells and hilar pyramidal neurons expressed approximately equal levels of α1 and α3. In the cerebellum, Purkinje and Golgi cells were rich in α3 mRNA, while the granule cells appeared to express only α1 transcripts. The distribution of functional sodium pump protein, as localized by [ 3H]ouabain binding, was highest in the neuropil of the hippocampus and cerebral cortex, and lowest over perikarya and white matter. [ 3H]ouabain did not bind to α1 pump units, as confirmed by the complete absence of labeling over the choroid plexus, a tissue expressing only α1 mRNA. In the cerebellum, regions of dense [ 3H]ouabain binding were localized to the granule cell layer, the inner third of the molecular layer in the basket region, and the deep cerebellar nuclei. Surprisingly, the dense neuropil in the outer 2 3 of the molecular layer lacked high [ 3H]ouabain binding. Thus, functional α3 sodium pump units appear distributed to the axon terminals and not to apical dendrites of Purkinje, Golgi and basket cells. A similar pattern of increased [ 3H]ouabain binding in axonal but not dendritic fields of α3-enriched neurons was present in the cerebral cortex and the hippocampus. Considering that many α3-enriched neurons are of the Golgi I type with long axons, the α3 isoform may be preferentially directed into axons to function in presynaptic membranes.