Rat Brain: Distribution of Immunoreactivity of PE‐11, a Peptide Derived from Chromogranin B

Abstract

An antiserum was raised against the peptide PE-11 whose sequence is present in the chromogranin B molecule. The antiserum reacts only with the free C-terminal end of this peptide. PE-11 immunoreactivity in brain was characterized by molecular size exclusion high performance liquid chromatography. Only the free peptide and a N-terminally elongated peptide were detected, indicating that proteolytic processing of chromagranin B in brain is quite extensive. In immunohistochemistry PE-11 immunoreactivity was found in varicosities, fibres and perikarya throughout the brain. Strong staining was detected in the shell sector of the nucleus accumbens, in the lateral septum, in subregions of the extended amygdala, in some areas of the hippocampus and of the hypothalamus, in the locus coeruleus, in the Purkinje cells of the cerebellum and in the dorsal horn of the spinal cord. Our results, which demonstrate significant processing of chromogranin B in brain and its widespread distribution, can be taken as an indication that chromogranin B represents a precursor of peptides with functional relevance for this organ.

Beta cell chromogranin B is partially segregated in distinct granules and can be released separately from insulin in response to stimulation

“…The polyclonal antibodies (pAb) were a guinea pig antiinsulin (DAKO, Milan, Italy) and two rabbit anti-CgBs, gifts of P. Rosa (Consiglio Nazionale delle Ricerche, Milan, Italy) and A. Laslop (PE-11; University of Innsbruck, Austria) [26,27]. The mouse monoclonal antibodies (mAb) were an IgG2a anti-rat CgB (CIRO) from our laboratory [28]; an anti-insulin (Sigma-Aldrich, Milan, Italy) and an anti-human CgB (67-C7-2), gift of W. B. Huttner (Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany); an anti-trans-Golgi network (TGN) marker (TGN38) and an anti-GM130 (Golgi complex marker; BD Biosciences, San Jose, CA, USA); an antilysosomal-associated membrane protein 1 (LAMP1) (LYC16; Calbiochem, Darmstadt, Germany); two antiprotein tyrosine phosphatase-like protein-2 (ICA512/IA-2), mICA512cyto, gift of M. Solimena (Technical University of Dresden, Germany) [29] and 76F from our laboratory [30].…”

Further Evidence that Behavioral Tests and Neuropeptide mRNA and Tissue Level Alterations Can Differentiate between Typical and Atypical Antipsychotic Drugs

“…Specific radioimmunoassays for enkephalin, chromogranin B (PE-11) (Kroesen et al 1996), and secretogranin II (secretoneurin) were performed as previously described (Marksteiner et al 1993a) with slight modifications. Briefly, samples and standards were incubated with antiserum for 24 hours.…”

“…The generation of the PE-11 antiserum was described previously (Kroesen et al 1996). Briefly, it was generated against a synthetic peptide (PE-11) corresponding to rat chromogranin B 552-562 (Forss et al 1989) which was identical to the human sequence.…”

“…They are endoproteolytically processed to smaller peptides, and are released after neuronal stimulation (Huttner et al 1991). The distribution of chromogranin B (Mahata et al 1991;Kroesen et al 1996), secretogranin II mRNA, and their immunoreactivity (Marksteiner et al 1993a) have been studied in rat brains. The biosynthesis of chromogranins is regulated by different stimuli (Shen and Gundlach 1996).…”

“…The dorsolateral caudate putamen and prefrontal cortex were dissected according to the Atlas of Paxinos: (Paxinos and Watson 1986). The extraction procedure was performed as previously described (Marksteiner and Sperk 1988).Specific radioimmunoassays for enkephalin, chromogranin B (PE-11) (Kroesen et al 1996), and secretogranin II (secretoneurin) were performed as previously described (Marksteiner et al 1993a) with slight modifications. Briefly, samples and standards were incubated with antiserum for 24 hours.…”