Immunochemical detection of diverse molecular species of horseradish peroxidase recovered from rat superior cervical ganglion following retrograde transport

Abstract

The sympathetic innervation of the submandibular gland (SMG) by neurons of the superior cervical ganglion (SCG) was used as a model system to characterize the different molecular species of retrogradely transported horseradish peroxidase (HRP). Following injections of 3 mg of HRP into the SMG, enzymatically active HRP was demonstrated by histochemistry in SCG neurons for up to 14 days. Parallel immunochemical studies were conducted on electroblots of SCG homogenates from other rats subjected to similar SMG injections of HRP. This technique allowed the detection of nanogram amounts of HRP. These experiments demonstrated that an immunochemically detectable 40 K d species of HRP was retrogradely transported and retained in SCG neurons for up to 6 days. Beyond 8 days, only lower molecular weight (15–30 K d) immunoreactive breakdown products of HRP were observed in SCG homogenates. Similar spontaneous HRP breakdown products displayed enzymatic activity in non-denaturing gels. These studies suggest that retrogradely transported HRP is degraded by neurons to lower molecular weight species which retain enzymatic activity. We conclude that morphological studies of retrogradely transported HRP visualized by histochemical methods reflect the response of neurons to a variety of enzymatically active molecular forms of HRP.