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The MDR1 gene product, P-glycoprotein (P-gp), was shown to confer multidrug resistance to cancer cells, but its overexpression is also suggested to be involved in pharmacoresistance of epilepsy by acting as an energy-dependent drug-efflux pump in the blood-brain barrier (BBB). In normal brain tissue, P-gp is almost exclusively expressed by capillary endothelial cells (EC) of the BBB, whereas little or no expression is detected in other cell types. Increased P-gp expression was observed after seizures, but localization of this increase, i.e., within brain capillary EC or within parenchymal or perivascular astrocytes, which contribute to the BBB function, is controversial. To test whether these antithetic data arise from unusual properties of the antigen itself, we compared different immunohistochemical techniques and monoclonal or polyclonal antibodies to P-gp in normal rat brain and rat brain after kainate-induced seizures. Using acetone-fixed cryostat sections of snap-frozen tissue, strong P-gp labeling was detected in EC and, after seizures, in hippocampal neurons, but not in astrocytes. In contrast, EC and neuronal P-gp immunolabeling were not seen in paraformaldehyde-fixed sections, whereas both perivascular and parenchymal astrocytes exhibited strong P-gp labeling after seizures. The lack of P-gp labeling in EC by paraformaldehyde fixation, was reversed by treatment of the sections with acetate/ethanol. These experiments demonstrate that various fixation conditions have a striking effect on the immunohistochemical localization of P-gp in rat brain and detection of its increased expression by seizures. When data obtained from different immunohistochemical techniques are taken together, seizures seem to induce overexpression of P-gp in four different cell types, i.e., EC, perivascular astrocytes, parenchymal astrocytes, and neurons.

journal of histochemistry and cytochemistry/the journal of histochemistry and cytochemistry

Immunohistochemical Localization of P-glycoprotein in Rat Brain and Detection of Its Increased Expression by Seizures Are Sensitive to Fixation and Staining Variables