Immunohistochemical Analysis of the α2 subunit of (Na+, K+)-ATPase in Mouse Brain

Janice, Michael J., Archana Wadhawan, and Mary Lou Caspers

The (Na+, K+)-ATPase enzyme catalyzes the active transport of Na+ and K+ across cell membranes and helps to reestablish ion balance after a neuron has fired.  It is composed of α and β subunits; three isoforms of the α subunit exist in brain.  Previous work in this laboratory, using [3H]ouabain, has detected the presence of the α2 and α3 subunits of the (Na+, K+)-ATPase in mouse brain sections (24 micron).  This laboratory has also used western blot analysis of homogenized tissue to determine the approximate concentration of the α2 subunit in the brain.  The western blot method is unable to detect the precise location of this subunit in brain tissue.  Therefore, work has begun using an indirect immunohistochemical procedure in which horseradish peroxidase luminescence is used in order to both quantitatively determine the concentration of the α2 subunit and ascertain the location of the subunit in brain sections.  Preliminary trials to date have used coronal sections in the rostral portion of the mouse brain.  The corpus callosum appears to have a higher concentration of the α2 subunit than any other region of the mouse brain which has been tested thus far.  More trials will need to be performed to analyze the caudal region of the brain and to reduce background luminescence. (We gratefully acknowledge the Department of Chemistry and Biochemistry, University of Detroit Mercy for support of this project.)