Regulation of Angiotensin II–induced Neuromodulation by MARCKS in Brain Neurons

Angiotensin II (Ang II) exerts chronic stimulatory actions on tyrosine hydroxylase (TH), dopamine β-hydroxylase (DβH), and the norepinephrine transporter (NET), in part, by influencing the transcription of their genes. These neuromodulatory actions of Ang II involve Ras-Raf-MAP kinase signal transduction pathways (Lu, D., H. Yang, and M.K. Raizada. 1997. J. Cell Biol. 135:1609–1617). In this study, we present evidence to demonstrate participation of another signaling pathway in these neuronal actions of Ang II. It involves activation of protein kinase C (PKC)β subtype and phosphorylation and redistribution of myristoylated alanine-rich C kinase substrate (MARCKS) in neurites. Ang II caused a dramatic redistribution of MARCKS from neuronal varicosities to neurites. This was accompanied by a time-dependent stimulation of its phosphorylation, that was mediated by the angiotensin type 1 receptor subtype (AT₁). Incubation of neurons with PKCβ subtype specific antisense oligonucleotide (AON) significantly attenuated both redistribution and phosphorylation of MARCKS. Furthermore, depletion of MARCKS by MARCKS-AON treatment of neurons resulted in a significant decrease in Ang II–stimulated accumulation of TH and DβH immunoreactivities and [³H]NE uptake activity in synaptosomes. In contrast, mRNA levels of TH, DβH, and NET were not influenced by MARKS-AON treatment. MARCKS pep^148–165, which contains PKC phosphorylation sites, inhibited Ang II stimulation of MARCKS phosphorylation and reduced the amount of TH, DβH, and [³H]NE uptake in neuronal synaptosomes. These observations demonstrate that phosphorylation of MARCKS by PKCβ and its redistribution from varicosities to neurites is important in Ang II–induced synaptic accumulation of TH, DβH, and NE. They suggest that a coordinated stimulation of transcription of TH, DβH, and NET, mediated by Ras-Raf-MAP kinase followed by their transport mediated by PKCβ-MARCKS pathway are key in persistent stimulation of Ang II's neuromodulatory actions.