ACTH, cyclic nucleotides, and brain protein phosphorylation in vitro

Abstract

Endogenous phosphorylation of proteins from rat brain synaptosomal plasma membranes was studied in vitro. Cyclic AMP (cAMP) markedly stimulated(32)P incorporation in three protein bands with molecular weights of 75,000, 57,000, and 54,000, respectively. The effect of the behaviorally active peptide ACTH1-24 on this endogenous phosphorylation in vitro was studied using peptide concentrations from 10(-10) to 10(-4) M. In a number of protein bands, a biphasic effect of ACTH1-24 was observed: in concentrations of 10(-4)-10(-5) M, a reduced amount of(32)P was found; in concentrations of 10(-6)-10(-7) M, hardly any effect could be detected, whereas consistently at concentrations around 10(-8) M, a significant decrease was again observed. The phosphoprotein bands affected by in vitro addition of ACTH1-24 were of a smaller molecular weight than those affected by in vitro addition of cAMP.

Historical review of research on protein kinase C in learning and memory

“…The ultimate goal of PKC activation, protein phosphorylation in a correct spatial-temporal fashion, has focused attention to the target substances. Since the discovery in the mid-1970s of a major PKC substrate participating in synaptic plasticity (Fl, also referred to as GAP43 or BSO; Routtenberg and Ehrlich, 1975;Zwiers et al, 1976) an impressive differentiation of over 100 enzyme targets and numerous physiological actions have been described (Liu, 1996;Casabona and Ramakers et al, …”

Pituitary hormones influence polyphosphoinositide metabolism in rat brain

“…The effects of peptide hormones on phosphoinositide metabolism are of special interest in view of the influence of ACTH and related peptides on brain membrane protein phosphorylation [9,10]. With respect to phosphorylation of membrane proteins, changes in membrane permeability in nerve and peripheral tissue have been reported (see [23]).…”

“…To prevent the rapid postmortem hydrolysis of diphosphoinositide and triphosphoinositide [ 111, the rats were killed by whole body immersion into liquid nitrogen for 10 s, followed by rapid dissection of the cold but not frozen brain, at 0-4°C. The limbic system (hippocampus, septum, basal ganglia, diencephalon, pyriform cortex) was dissected as in [ 121. Crude mitochondrial preparation was according to [9]. The limbic system was homogenized in 10 vol.…”

“…In view of these findings and our recent results on the regulatory role of peptide hormones and fragments of peptide hormones on synaptic plasma membrane protein phosphorylation [9,10], it was of interest to investigate whether such peptides influence brain membrane phospholipid metabolism.…”

α-MSH-induced changes in protein phosphorylation of Cloudman S91 mouse melanoma cells

“…Routinely, 10 pg were applied per slot. Electrophoresis was performed at 120 V/30 mA for 6-7 h. Proteins were stained with 0.1% Fast Green according to Zwiers et al (1976). Gels were dried under vacuum at 50 o C and subjected to autoradiography.…”