DE-NOVO SYNTHESIS OF GAP-43 - IN-SITU HYBRIDIZATION HISTOCHEMISTRY AND LIGHT AND ELECTRON-MICROSCOPY IMMUNOCYTOCHEMICAL STUDIES IN REGENERATING MOTOR-NEURONS OF CRANIAL NERVE NUCLEI IN THE RAT-BRAIN

In order to investigate the modulation of the synthesis and the subcel lular localization of the growth associated protein GAP-43 in neuronal cell bodies we have taken advantage of the well known regenerative pr operties of axotomized motor neurons of the facial and hypoglossal nuc lei. Alterations in the levels of GAP-43 mRNA containing cells were st udied by in situ hybridization histochemistry. The protein localizatio n was examined using immunohistochemistry at the light and electron mi croscopic levels. Neurons from the control side showed undetectable le vels of both GAP-43-like immunoreactivity and GAP-43 mRNA levels. Wher eas axotomized neurons exhibited a marked increase in GAP-43 mRNA leve ls and in GAP-43-like immunoreactivity. Three to 50 days after axotomy , motor neurons ipsilateral to the lesion displayed a dense reticular or filamentous perinuclear distribution of the immunoreactivity in som ata and proximal dendritic processes, corresponding to the location of the Golgi apparatus in these neurons. At the electron microscopic lev el the immunoreactivity was located in the cisternae of the Golgi comp lex and found to be associated with trans-side vesicles of these compl exes. The myelinated fibers of the transectomized facial nerve also pr esented an intense GAP-43-like immunoreactivity. Twenty-one days after the axotomy a decay in the number of immunostained neurons and in the intensity of immunolabeled somata was observed. Our study reveals a r apid induction of GAP-43 mRNA and protein after axotomy. The localizat ion of the newly synthesized GAP-43-like immunoreactivity to the Golgi apparatus seen in the present work suggests an early association of t his protein with newly formed membranes prior to transport toward the terminals through the axons.