Time course of action of pertussis toxin to block the inhibition of stimulated insulin release by norepinephrine.

Abstract

Male Sprague-Dawley rats were injected ip with 1 microgram pertussis toxin (PTX)/100 g BW. The rats were killed 24, 48, and 72 h after injection, and their pancreases were removed. At each time point, insulin secretion by isolated islets was measured under basal and glucose-stimulated conditions and in the absence and presence of norepinephrine. cAMP levels were measured under basal and forskolin-stimulated conditions in the absence and presence of norepinephrine. PTX-induced ADP ribosylation of Gi/Go proteins in vivo was monitored by ADP ribosylation in vitro using PTX and 32P-labeled NAD and also by Western blotting. At 24 h, 1 microM norepinephrine inhibited glucose-stimulated insulin secretion by 92% in the control islets, but by only 53% in the PTX-treated islets; at 48 h, norepinephrine still inhibited secretion (by 40%) in the PTX-treated islets; at 72 h, the inhibitory effect of norepinephrine was abolished. Therefore, contrary to recent suggestions, all of the effect of norepinephrine to inhibit insulin release is PTX sensitive. The effects of PTX on the ability of norepinephrine to lower cAMP levels were similar to those observed for the inhibition of insulin release. PTX partially blocked the effect of norepinephrine to lower cAMP levels at 24 and 48 h, and the block was complete after 72 h. The extent of the in vivo ADP ribosylation of the Gi/Go proteins, monitored at each time point by in vitro [32P]ADP-ribosylation and Western blotting, demonstrated a profound ADP ribosylation at 48 and 72 h. As detected by Western blotting, the rates of ADP ribosylation by PTX and the onset of decreased expression varied among the different G-proteins. G alpha o was virtually eliminated after only 24 h of PTX treatment. G alpha i2 was markedly affected by 48 h; G alpha i3 was little affected until 72 h.

Masked excitatory action of noradrenaline on rat islet ?-cells via activation of phospholipase C

“…These findings indicate that the inhibitory effect of NE on insulin secretion is mediated by a PTX-sensitive pathway, and that an excitatory pathway for NE to stimulate insulin secretion is also present in islets. The present results with PTX treatment differ in part from those of other studies [22], in which treatment with PTX abolished only the inhibitory effect of NE. The reason for this discrepancy is not clear, but the efficacy of PTX may differ with different routes of application.…”

“…In the study cited PTX was given by i.p. injection [22], whilst we applied it directly to the islets or single cells. Over-night treatment of islets with direct application of PTX is known to inhibit the G i proteinlinked pathway [11].…”

“…This inhibitory process involves α 2 -adrenoceptors, leading to activation of pertussis toxin (PTX)-sensitive G i proteins and inhibition of adenylate cyclase [11,22]. Coupling of G i proteins, inhibition of adenylate cyclase and the subsequent inhibition of cAMP production has been investigated extensively using galanin, which is contained in and released from the sympathetic nerve terminal [9] and binds to its G i protein-linked receptor [7,24].…”

Pertussis Toxin Administered Spinally Induces a Hypoglycemic Effect on Normal and Diabetic Mice

“…For example, systemically injected PTX produces a hypoglycemic effect [4,5]. Furthermore, an in vitro study by Komatsu et al [7] reported that PTX appears to increase the secretion of insulin. PTX causes hypoglycemia due to enhancement of insulin secretion, leading to the original denomination of the toxin as ‘islet-activating protein' by Ui [22].…”

“…For example, systemically administered PTX produces a hypoglycemic effect [5,6]. Furthermore, in an in vitro study, PTX appeared to increase the secretion of insulin [7]. Also, several cell biological processes in insulin-producing cells are reversed or counteracted by PTX [8].…”