R.E. Catalán

Substance P stimulates translocation of protein kinase C in brain microvessels

The action of substance P on protein kinase C in cerebral microvessels, isolated from bovine, was investigated. We found that in untreated microvessels 85% of the total activity was localized to the cytosolic fraction. Substance P caused a shift of activity to the membrane fraction, accompanied by a loss of activity in the cytosolic fraction. This effect resulted in dose-dependence and it was evident after 5 min treatment. These data suggest that substance P may be involved in the regulation of processes underlying protein phosphorylation in the blood-brain barrier.

Insulin action on brain microvessels; Effect on alkaline phosphatase

The effects of insulin on brain alkaline phosphatase activity have been examined. Insulin inhibited the activity of alkaline phosphatase on brain microvessels in in vitro experiments. The inhibition observed was of the non-competitive type. These observations indicate that the hormone is able to induce neurochemical modifications revealed in this case as changes in the phosphate transfer enzymes in brain microvessels.

PAF-induced activation of polyphosphoinositide-hydrolyzing phospholipase C in cerebral cortex

The action of platelet-activating factor (PAF) on phosphoinositide hydrolysis was studied in rat brain slices. PAF produced a significant increase of 32P incorporation into phosphoinositides and phosphatidic acid (PA), in a dose- and time-dependent manner. Concomitantly, an increase of inositol phosphates and diacylglycerol (DAG) production was observed. Both inositol bisphosphate (IP2) and inositol trisphosphate (IP3) were detected as early as 5 s and they returned immediately to basal levels; concomitantly, formation of inositol monophosphate (IP) was detected. These findings demonstrated that PAF causes a rapid hydrolysis of polyphosphoinositides in cerebral cortex by a phospholipase C-dependent mechanism followed by subsequent resynthesis.

Evidence for a role of somatostatin in lipid metabolism of liver and adipose tissue

We have studied the effects of somatostatin on lipid metabolism in liver and adipose tissue of fasted mice. The animals were injected subcutaneously with 8 micrograms somatostatin and killed 5 min after injection. In vivo incorporation of [14C]acetate into triglycerides in both tissues and into hepatic cholesterol was significantly enhanced by somatostatin. Concomitantly, a decrease of triglyceride lipase activity was observed, which corresponds well with the variation undergone by cyclic AMP-protein kinase system. In addition, a marked increase of serum cholesterol levels was observed. Additionally, in vitro experiments were also performed by employing 2.4 X 10(-6) M somatostatin. The results showed that the direct effect of somatostatin on liver seems to be a decrease in acetate uptake. The results obtained with the adipose tissue were similar to those obtained in in vivo conditions. On the other hand, when somatostatin was administered in vivo, the ability to incorporate ortho[32P]phosphate into phospholipids was enhanced in both tissues. Likewise in the in vitro experiments with [14C]acetate, the somatostatin seems to act by decreasing the ortho[32 P]phosphate uptake in liver. While in adipose tissue the somatostatin only caused a strong increase in the specific activity of phosphatidylcholine. These data demonstrate in fasted mice that somatostatin is able to counteract the lipolytic manifestations of the fasted state.

SELECTIVE TIME-DEPENDENT EFFECTS OF INSULIN ON BRAIN PHOSPHOINOSITIDE METABOLISM

The effect of insulin on phosphoinositide metabolism in the cerebral cortex was examined using 32P as precursor. A maximal increase was detected as early as 15 s; phospholipid labeling declined after this initial peak but then increased to another maximum at 30 min. The levels of these phospholipids were unchanged at the earliest time examined, but at 30 min insulin caused an increase in the content of all phospholipids tested. In pulse-chase experiments, insulin stimulated depletion of 32P-labeled phosphoinositides only at 15 s. On the other hand, insulin treatment caused a biphasic diacyglycerol (DAG) production. We conclude that in cerebral cortex, insulin has a dual mechanism of action on phosphoinositide metabolism. First, insulin causes a rapid but transient hydrolysis of phosphoinositides by a phospholipase C-dependent mechanism, followed by subsequent resynthesis; thereafter, insulin increases de novo phospholipid synthesis.

Alterations in rat lipid metabolism following ecdysterone treatment

The influence of ecdysterone on the lipid metabolism in liver and adipose tissue from rat was investigated using 14C-acetate and 32P-orthophosphate as precursors. Ecdysterone produced an increase in 14C-acetate incorporation into triglycerides. A concomitant decrease in free fatty acids and diglycerides was observed. The effect of ecdysterone on triglyceride lipase activity was investigated and a significant decrease was found. Ecdysterone produced a significant increase in the specific activity of phosphatidylethanolamine and phosphatidylserine in liver. On the contrary, the specific activity of phosphatidylcholine was reduced. In adipose tissue, the most evident effect observed was the increase of specific activity of phosphatidylcholine. These results contribute to knowledge of the heterophylic action of ecdysterone.

Ecdysterone induces acetylcholinesterase in mammalian brain

The effects of ecdysterone on brain acetylcholinesterase (AChE) in immature and adult rats of both sexes have been studied in in vitro conditions. Ecdysterone produced an increase of AChE in rat brain slices. The most remarkable effect was found in immature male rats. In vitro assay using a purified AChE from electric eel showed no effect. Pretreatment with cycloheximide or actinomycin D abolished the ecdysterone action on brain AChE. These results support the idea that induction of AChE may be involved in the heterophilic action of ecdysterone.

Evidence for a regulatory action of vanadate on protein phosphorylation in brain microvessels

We investigated the action of vanadate on protein phosphorylation in microvessels isolated from rat brain. We found that a stimulation of protein phosphorylation from 32P-ATP occurs, in the presence of different concentrations of vanadate, 10(-3) M being the most effective dose. This action was time-dependent, and it was more evident after 60 s of treatment. The contribution of ATPase inhibition caused by vanadate appears to be negligible. In addition a stimulation of cAMP-dependent protein kinase activity was observed. The pattern of protein phosphorylation showed that exposure to 10(-3) M vanadate resulted in a nonspecific stimulation of protein phosphorylation concomitantly with a selective inhibition of the 55 KDa protein phosphorylation. The nature of this protein is also discussed.

Substance P induces alterations on cerebral lipids involved in membrane fluidity

This study was undertaken to examine the variations in rat brain of cholesterol, phospholipid and phospholipid fatty acid composition induced by substance P. The cholesterol content was increased by substance P; concomitantly, an increase of the ratio cholesterol/phospholipid was observed. These changes do not appear to be responsible of the stimulation observed in Na+,K+-ATPase activity by substance P action. Phospholipid fatty acid analysis revealed that the peptide induced a decrease in both linoleic and arachidonic acids content.

Inhibition of cyclic AMP-dependent protein kinase by somatostatin in slices of mouse brain: dependence on extracellular calcium.

Over a concentration range from 2 to 8 micrograms/ml, cyclic somatostatin inhibited cyclic AMP-dependent protein kinase and cyclic AMP binding activity in slices of mouse brain. An inverse pattern of variation was observed in the activity of phosphoprotein phosphatase. The action on kinase activity was observed in the presence of physiological concentrations of extracellular calcium, but when the concentration of calcium was increased the inhibition of kinase was not observed. These results support the idea that the protein kinase system may be involved in the action of somatostatin in the central nervous system and they are consistent with the hypothesis that somatostatin affects calcium flux.