T. Florio

Somatostatin inhibition of adenylate cyclase activity in different brain areas

Somatostatin receptors have been identified in different brain areas but the characterization of their postreceptor effect is still lacking. In this study we analyze the somatostatin effect on adenylate cyclase activity in different brain regions, namely frontal cortex, striatum, hypothalamus and hippocampus. Somatostatin inhibited basal adenylate cyclase activity in all brain areas, being maximally effective in the frontal cortex (-42%). Moreover, somatostatin inhibited both dopamine- and norepinephrine-stimulated adenylate cyclase activity in the examined cerebral regions showing a higher effectiveness than in basal conditions. VIP stimulation of adenylate cyclase was also reduced by somatostatin. The peptide inhibited by 50% forskolin-stimulated (10 nM to 10 microM) enzyme activity in frontal cortex and hypothalamus (in hippocampus the inhibition reached only -25%) showing a non-competitive pattern of inhibition. Finally, pertussis toxin pretreatment abolished the somatostatin inhibition of forskolin-stimulated frontal cortex adenylate cyclase activity. These results show that brain somatostatin receptors are coupled in an inhibitory way with adenylate cyclase enzyme that may represent one of the postreceptor mechanisms mediating the somatostatin modulation of brain functions.

Somatostatin and SMS 201-995 reverse the impairment of cognitive functions induced by cysteamine depletion of brain somatostatin

The involvement of somatostatin in the organization of cognitive functions was studied. We assessed changes in learning and memory processes by studying the effects of cysteamine, a compound that decreases somatostatin-like immunoreactivity in the brain, somatostatin and the potent somatostatin analogue, SMS 201-995, on active avoidance behaviour, assessed with a shuttle box apparatus, or on passive avoidance behaviour. Cysteamine induced a loss of the conditioned active avoidance response acquired after 3 weeks of daily trials. The effect was observed 2 h (-29%) and 4 h (-51%) after cysteamine treatment (300 mg/kg s.c.) and disappeared after 24 h. Intracerebroventricular administration of somatostatin or SMS 201-995 to cysteamine-treated rats significantly reversed the cysteamine effects on the conditioned avoidance responses. Similar results were obtained on passive avoidance behaviour. We also investigated the effect of cysteamine treatment on brain somatostatin-sensitive adenylate cyclase. We observed that adenylate cyclase activity in the frontal cortex of cysteamine-pretreated animals was more sensitive to inhibition by the SRIF analogue, SMS 201-995, than it was in control animals. This effect was observed at concentrations of SMS 201-995 that were ineffective in control tissue. These results show that disruption of somatostatinergic transmission affects cognitive functions of rats.

Somatostatin inhibition of anterior pituitary adenylate cyclase activity: different sensitivity between male and female rats

Somatostatin (SRIF) is a potent inhibitor of growth hormone (GH) secretion. Although cyclic AMP (cAMP) has been suggested as intracellular mediator of SRIF action, a complete characterization of its effect and the different sensitivity between male and female animals, has not yet been carried out. In this study SRIF inhibited basal and GH-releasing factor (GRF) stimulated anterior pituitary adenylate cyclase activity with a greater effectiveness in male than in female glands. Similarly SRIF reduction of forskolin-stimulated anterior pituitary adenylate cyclase activity, was more pronounced in male than in female animals. By using pertussis toxin (PTX), which uncouples inhibitory receptors from adenylate cyclase catalytic subunit, SRIF inhibition of both basal and forskolin-stimulated adenylate cyclase activity was nearly abolished. These results show that anterior pituitary SRIF receptors are coupled in an inhibitory fashion with adenylate cyclase enzyme, and that male rat adenohypophyses are more responsive to SRIF inhibition.

Age-related alterations of somatostatin gene expression in different rat brain areas.

Numerous experimental and clinical studies have demonstrated that brain somatostatinergic neurotransmission plays an important role in the modulation of several brain functions, including learning and memory processes. Due to the gradual decline of cognitive performances occurring during aging, we evaluated whether an age-related modification of brain somatostatin gene activity occurred in discrete rat brain areas. Our study demonstrates that a significant reduction of pre-prosomatostatin mRNA levels occurred in aged animals (25 months) in the frontal cortex (-49%), in the parietal cortex (-80%) and in the striatum (-69%), despite the absence of changes in beta-actin gene expression. Conversely, no statistical differences were observed in the pre-prosomatostatin mRNA content of old animals in the hypothalamus. These results demonstrate that age-related alterations in somatostatin gene expression occur in the rat, and suggest that such alterations may be involved in the behavioral and cognitive impairments that occur during the aging process.

Effect of interleukin 1 beta on transducing mechanisms in 235-1 clonal pituitary cells: Part I: Modulation of adenylate cyclase activity

In the present study we evaluated the effect of interleukin 1-beta on adenylate cyclase activity in 235-1 pituitary cell line. The dose-response curve of interleukin 1 beta effect on adenylate cyclase activity showed a significant inhibition of basal enzyme activity at 1 pM concentration, while the inhibition of forskolin stimulated adenylate cyclase activity was more pronounced and evident at both 0.01 and 1 pM concentrations. The action of the monokine on basal enzyme activity was almost completely reverted by polyclonal anti interleukin 1 beta antibody. The incubation of the cells for 48h with interleukin 1 beta showed a different pattern of response. The inhibitory effect of interleukin 1 beta on adenylate cyclase activity disappeared, while the highest concentration of interleukin 1 beta tested, caused a meaningful stimulation of adenylate cyclase activity which is not present in acute condition. These data show that interleukin 1-beta interacts with the cAMP-generating system in the 235-1 clonal pituitary cells.

Pertussis toxin pretreatment abolishes dihydropyridine inhibition of calcium flux in the 235-1 pituitary cell line

In the present study we used 235-1 cells, a prolactin secreting clone derived from a pituitary tumor. In these cells maitotoxin, a calcium channels activator, likely acting on voltage sensitive calcium channels, increases intracellular free calcium measured by Quin 2 technique. Maitotoxin stimulation of calcium flux was inhibited both by nicardipine and verapamil in a dose dependent manner. Pertussis toxin pretreatment does not modify maitotoxin activation of calcium channels, while completely abolishes nicardipine inhibition of maitotoxin induced voltage sensitive calcium channels activation, without affecting verapamil effect. These results suggest a possible involvement of a pertussis toxin sensitive G protein in dihydropyridine inhibition of voltage sensitive calcium channels.

Dihydropyridine Modulation of Voltage‐Activated Calcium Channels in PC12 Cells: Effect of Pertussis Toxin Pretreatment

Abstract: In this study, we report the effect of pertussis toxin pretreatment on dihydropyridine modulation of voltage‐sensitive calcium channels in PC12 cells. The rise in intracellular calcium concentration caused by potassium depolarization is not affected significantly by pertussis toxin pretreatment. Nicardipine, a dihydropyridine derivative, added either before or after potassium‐induced depolarization, reduces the resultant elevation in cytosolic calcium level both in control and in pertussis toxin‐treated cells. The dihydropyridine agonist Bay K 8644, when added before potassium, is able to enhance the potassium‐induced spike of cytosolic calcium levels, an effect significantly reduced by pertussis toxin pretreatment. Moreover, the addition of Bay K 8644 after potassium holds the intracellular calcium concentration at a cytosolic sustained level during the slow inactivating phase of depolarization. This effect of Bay K 8644 is inhibited by nicardipine. Pertussis toxin pretreatment slightly weakens the effect of Bay K 8644 when added after potassium‐induced depolarization, whereas it significantly reduces the nicardipine inhibition of cytosolic calcium rise stimulated by potassium and Bay K 8644, but not by potassium alone. In conclusion, our findings suggest that a pertussis toxin‐sensitive guanine nucleotide regulatory protein could be involved in the interaction between dihydropyridine derivatives and voltage‐dependent calcium channels.

Interleukin 1 beta inhibition of TRH-stimulated prolactin secretion and phosphoinositides metabolism

The effect of interleukin 1 beta on prolactin secretion and on phosphoinositide turnover in anterior pituitary cells was evaluated. Interleukin 1 beta significantly inhibited TRH-stimulated prolactin secretion assessed by the reverse hemolytic plaque assay. In particular, the cytokine reduced the percentage of plaque forming cells, the plaque mean area, the large plaques percentage. TRH-stimulated inositol phosphate production was also significantly inhibited by interleukin 1 beta. This study shows that interleukin 1 beta reduces TRH-induced prolactin secretion through a direct action on pituitary cell, and attenuates the TRH-stimulated phosphoinositide breakdown. This latter effect may suggest that the reduced lactotropes sensitivity to TRH action may be partially due to interleukin 1 beta inhibition of phosphatidylinositol breakdown.

Fipexide improvement of cognitive functions in rat: Behavioural and neurochemical studies

Dopamine neurotransmission plays a role in learning, memory and related cognitive processes. We evaluated the effect of a nootropic drug, fipexide, a parachloro-phenossiacetic acid derivative, both on behavioural and biochemical parameters. The compound was able to cause an improved performance on active avoidance test, when administered just before the trial, being also effective on memory retention and recall experiments. Fipexide was also able to revert the impairment of the acquired behaviour caused by sulpiride in pretrained rats, while it failed to be effective on the haloperidol deconditioning effect. Striatal adenylate cyclase activity, from fipexide pretreated rats, was significantly reduced both in basal and dopamine stimulated conditions. Furthermore, fipexide, directly added to membrane preparations, was able to inhibit striatal adenylate cyclase activity. These results, along with our previous reports on fipexide effect on locomotor activity, allow us to hypothesize that the positive effect of this drug on cognitive performance seems to be mediated, at least partially, by dopaminergic neurotransmission.

Somatostatin and Brain Aging

In the past decade a rapid progress has been achieved in the isolation and characterization of a number of bioactive peptides from the brain, mostly from the hypothalamus, and the gastrointestinal tract. These peptides are known to influence and regulate many physiological processes in a variety of animal and human tissues. One of the peptides, that has received increasing attention in recent years, is somatostatin.