Effects of dietary cholesterol on pyroglutamyl aminopeptidase activity in mouse frontal cortex, pituitary, and adrenal glands.

Pyroglutamyl aminopeptidase (pGluAP) is an omega peptidase that hydrolyzes biologically active peptides, such as thyrotropin-releasing hormone (TRH), with neuronal and extraendocrine functions. We analyzed the effects of a cholesterol-enriched diet on soluble and membrane-bound pGluAP activity in frontal cortex, pituitary and adrenal glands of male and female mice using fluorimetric assays. Significant increases were observed in soluble pGluAP activity in the frontal cortex and adrenal glands in males and in the pituitary in females. Membrane-bound pGluAP activity was increased in the frontal cortex and pituitary of males and females after the mice were fed a cholesterol-enriched diet. These increases may produce changes in the metabolism of endogenous substrates, including TRH, which may be related to alterations in its neuromodulator functions and to the possible relationship between TRH and other neurotransmitter systems.

Calcium-dependent modulation by ethanol of mouse synaptosomal pyroglutamyl aminopeptidase activity under basal and K(+)-stimulated conditions.

We studied the in vitro effects of ethanol (25, 50 and 100 mM) on pyroglutamyl aminopeptidase activity (pGluAP), which has been reported as thyrotrophin-releasing-hormone-degrading activity. pGluAP was measured in presence or absence of calcium, under basal and K(+)-stimulated conditions, in synaptosomes and their incubation supernatant, using pyroglutamyl-beta-naphthylamide as substrate. In basal conditions, in synaptosomes, pGluAP was inhibited by ethanol in a calcium-independent way. In the supernatant, the response differed depending on the concentration of ethanol. Depolarization with K(+) modified pGluAP in synaptosomes and supernatant depending on the presence or not of calcium. In synaptosomes, in absence of calcium, the activity was inhibited at the highest concentrations of ethanol. In contrast, in the supernatant, under depolarizing conditions, ethanol increases pGluAP in absence of calcium. These changes may be in part responsible of the behavioural changes associated to alcohol intake.

[Aminoglycoside antibiotics neomycin and kanamycin inhibit the increase of of pyroglutamyl aminopeptidase activity by depolarizing synaptosomes of the frontal cortex of the rate]. / Los antibióticos aminoglucósidos neomicina y kanamicina inhiben el incremento de actividad piroglutamato aminopeptidasa obtenido tras despolarización de sinaptosomas de corteza frontal de rata.

INTRODUCTION: Thyrotrophin releasing hormone (TRH) has emerging in the last few years as a neuropeptide with important functions, not only as neurohormone into the hypothalamus-pituitary axis, but as neurotransmitter in several areas of the nervous system. Although little is known about its extra-endocrine functions, TRH has been related with several types of psychiatric disorders. Pyroglutamyl aminopeptidase (pGluAP) is the enzyme involved in the degradation of TRH. OBJECTIVES: The present research studies the levels of pGluAP activity under basal (resting) and KCl-stimulated (depolarized) conditions. The role of intracellular free calcium homeostasis, by means of the aminoglycoside antibiotics neomycin and kanamycin as voltage-dependent calcium channels blockers, is also studied. MATERIAL AND METHODS: Both pGluAP activity and intracellular free calcium concentration were analyzed in synaptosomes obtained from the frontal cortex of rats. Synaptosomes were incubated in artificial cerebrospinal fluid, under basal (resting) or KCl-stimulated (depolarized) conditions, with of without neomycin or kanamycin at different concentrations. RESULTS: Depolarization increases significantly pGluAP activity, which is completely abolished by neomycin and kanamycin at the lower concentrations used. On the contrary, aminoglycoside antibiotics do not block completely the increase on intracellular free calcium concentration induced by depolarization. Under basal conditions, no changes were found on pGluAP activity nor intracellular free calcium. CONCLUSIONS: pGluAP activity could regulate the neurotransmitter/neuromodulatory functions of TRH trough intracellular free calcium movements through aminoglycoside-sensitive voltage-dependent calcium channels. A role for inositol 4,5-bisphosphate breakdown products is also suggested.

Los antibióticos aminoglucósidos neomicina y kanamicina inhiben el incremento de actividad piroglutamato aminopeptidasa obtenido tras despolarización de sinaptosomas de corteza frontal de rata / Depolarization increases pyroglutamyl aminopeptidase activity in rat frontal cortex synaptosomes. Inhibitory effect of aminoglycoside antibiotics neomycin and kanamycin

Introducción. Uno de los neuropéptidos que está alcanzando un gran interés en los últimos años es la tirotropina o TRH, tanto por su papel como neurohormona en el eje hipotálamo-hipofisiario, como por su poco conocida función extra-endocrina. La acción neurotransmisora/neuromoduladora de la TRH es finalizada por la enzima piroglutamato aminopeptidasa (pGluAP). Objetivos. El presente trabajo se ha diseñado para estudiar el comportamiento de la actividad pGluAP en una situación basal o de reposo y tras la despolarización con KCl 50 mM. Para comprobar su posible relación con los niveles de calcio libre citosólico, se han utilizado los antibióticos aminoglucósidos neomicina y kanamicina como bloqueantes de canales de calcio dependientes de voltaje. Material y métodos. La actividad pGluAP, así como los niveles de calcio libre citosólico, se determinaron en sinaptosomas de corteza frontal de rata. Para ello se mantuvieron los sinaptosomas tanto en condiciones basales como despolarizantes, en un medio de incubación en presencia o ausencia de neomicina y kanamicina a diferentes concentraciones. Resultados. La despolarización de los sinaptosomas provoca un incremento altamente significativo de la actividad pGluAP, el cual es inhibido completamente por neomicina y kanamicina. Sin embargo, estos antibióticos aminoglucósidos no bloquean por completo el incremento en los niveles de calcio libre citosólico inducido por la despolarización. Ninguno de los parámetros estudiados se modifica en condiciones basales. Conclusiones. La actividad pGluAP puede controlar la función neurotransmisora/neuromoduladora de la TRH a través de los movimientos de calcio libre citosólico originados a través de canales de calcio sensibles a antibióticos aminoglucósidos, si bien hay que considerar el posible papel de los mensajeros intracelulares derivados del inositol 4,5-bisfosfato (AU)

[In vitro study of the effect of ethanol on pyroglutamyl aminopeptidase activity in mouse synaptosomes under basal and stimulated conditions]. / Estudio in vitro del efecto del etanol sobre la actividad piroglutamato aminopeptidasa en sinaptosomas de ratón en condiciones basales y despolarizantes.

INTRODUCTION AND OBJECTIVE: Pyroglutamyl aminopeptidase (pGluAP) is an omega peptidase which removes pyroglutamyl N-terminals residues from peptides and arylamidase derivatives. This activity is thought to be involved in the regulation of several physiological mechanisms on the central nervous system. pGluAP can modulate various susceptible endogenous substrates such as thyrotrophin-releasing hormone (TRH). It is well known that TRH plays an important role in the modulation of the behavioral changes induced by ethanol and others drugs. The aim of this work was to study the in vitro effects of ethanol (25, 50 and 100 mM) on the pGluAP activity and its ability for modulating the TRH. MATERIAL AND METHODS: pGluAP activity was measured in synaptosomes from cerebral cortex of mouse, using pyroglutamyl-beta-naphthylamide as substrate in basal and stimulated (K+ 25 mM) conditions, and in presence or absence of calcium on the buffer. RESULTS: In basal conditions, ethanol produced an inhibition of the pGluAP activity in presence or absence of calcium, being this inhibition non dose-related. However, the stimulation with K+ 25 mM did not produce a modification of pGluAP activity in presence of calcium, but produced a light increase in absence of it. Depolarization in presence or absence of calcium and ethanol produced an inhibition of pGluAP activity, which changed in function of the ethanol concentration used. CONCLUSIONS: Ethanol modifies pGluAP activity in basal conditions by a mechanism independent of calcium, but the changes observed after the stimulation with high K+ may be due to a calcium-dependent mechanism. These variations of pGluAP activity induced by ethanol, and their effects on their endogenous substrates, specially TRH, may be responsible for the behavioral changes associated to the alcoholism and mediated by TRH.

Mood disorders associated with acoustic neuromas.

OBJECTIVE: The primary purpose of this article is the presentation of three cases of manic and mixed states associated with an acoustic neuroma, and review of the literature on psychiatric symptoms accompanied by the tumor. METHODS: The cases were identified from 830 consecutive inpatient psychiatric admissions over age fifty-five years. Patients were assessed using a Structured Clinical Interview (SCID-R), and met DSM-III-R criteria for the diagnosis of bipolar disorder. The psychopathology seen in acoustic neuroma patients and the pathophysiologic mechanisms proposed to explain them are reviewed. RESULTS: The cases we report differ from other cases in the literature in that psychiatric symptoms began pre-operatively and remained for long periods post-operatively. The psychiatric signs and symptoms reported in acoustic neuroma patients are usually described as transient, and these include mood changes, agitation, persecutory delusions, hallucinations, and memory loss and confusional episodes. The disruption of brainstem structures including the auditory pathways, the cerebellum and the ascending reticular system may contribute to mood changes. Systematic studies are necessary to examine their relationship. Although psychological reactions attributable to surgery and facial paralysis may serve as contributory factors the evidence for their role was not striking in the cases we report.

Agitation and postdexamethasone cortisol levels in Alzheimer's disease.

The clinical correlates of hypothalamic-pituitary-adrenal (HPA) functioning were examined in 29 patients with probable Alzheimer's disease. The 8:00 a.m. postdexamethasone cortisol levels of these patients were highly correlated with higher agitation scores but not with the degree of depressed mood or memory impairment. The possible neural basis for the association between hypercortisolism and behavioral disturbance in Alzheimer's disease warrants further exploration and replication.

Restoration of cholinomimetic activity by clonidine in cholinergic plus noradrenergic lesioned rats.

The effects of combined lesions of forebrain cholinergic and noradrenergic systems on memory and responsivity to the memory enhancing effects of cholinomimetics were investigated in rats. Forebrain noradrenergic deficits produced by the injection of 6-hydroxydopamine into the ascending noradrenergic bundle (ANB) blocked the ability of cholinomimetics such as physostigmine and oxotremorine to enhance retention test performance in nucleus basalis of Meynert lesioned rats. Low doses of the noradrenergic receptor agonist clonidine, when administered in conjunction with cholinomimetics reversed this blockade. These results suggest that combined cholinergic/noradrenergic therapy may be of value in the treatment of some Alzheimer's disease patients.

Thyroid hormone action: early calorigenic effect on dispersed rat liver cells in the absence of protein synthesis.

Isolated dispersed rat liver cells were prepared by hypothyroid Sprague-Dawley rats. The cells were incubated under 95% O2/5% CO2 in Krebs-Ringer-bicarbonate buffer at pH 7.3-7.4 at 37 degrees C. The medium had been enriched with 2% bovine serum albumin (previously stripped of thyroid hormone) and 5-10 mM alanine as substrate. Two hour incubations were carried out with or without added triiodothyronine (T3) at 3 nM or 300-1,000 nM concentrations. Oxygen consumption determined at the end of the period of incubation with the Clark oxygen electrode showed stimulation above control values in the hormone treated flasks; parallel studies in which cycloheximide (100 microM) had been added to cells to block protein synthesis also showed enhanced oxygen consumption in response to T3. The results indicated a response to the hormone not dependent on new protein formation.

Cysteamine-induced depletion of central somatostatin-like immunoactivity: effects on behavior, learning, memory and brain neurochemistry.

The effects of a wide range of doses of systemically administered cysteamine were studied on locomotor behavior, passive avoidance memory, cortical and cerebrospinal fluid somatostatin-like immunoactivity and cortical levels of dopamine and norepinephrine. High doses of cysteamine (200 and 250 mg/kg s.c.) led to sustained locomotor activation. Doses of 150 mg/kg and above resulted in head and neck tremor and increased defecation. When cysteamine was administered immediately following the acquisition of a passive avoidance response, doses of 50 mg/kg and above resulted in significant attenuation of passive avoidance retention test performance. Cysteamine in doses of 50 mg/kg and above depleted cortical somatostatin-like immunoactivity by approximately 50%. The depletion of cortical somatostatin-like immunoactivity was accompanied by a rapid rise in somatostatin-like immunoactivity in cerebrospinal fluid. In addition to the depletion of somatostatin-like immunoactivity, high doses of cysteamine (150 mg/kg and above) produced changes in cortical levels of norepinephrine and dopamine, reminiscent of dopamine-beta-hydroxylase inhibition. The results of this series of experiments suggest that somatostatin, in addition to its effects on hormonal regulation, may play an important role in behavior and passive avoidance learning and memory. It is possible that the amnesia produced by cysteamine may have been due to the release of somatostatin into CSF from tissue stores, rather than somatostatin depletion per se. It is also possible that the catecholaminergic effects of high doses of cysteamine contribute to the behavioral deficits observed.

Animal models of Alzheimer's disease: behavior, pharmacology, transplants.

Physostigmine, oxotremorine, RS-86, and a combination of piracetam and lecithin, have all been studied in patients with Alzheimer's disease. Intravenous physostigmine produced a significant improvement in patients' ability to recognize words and particularly to distinguish words they had never seen before from words previously presented. A subgroup of Alzheimer's patients had a clinically meaningful improvement to treatment with oral physostigmine, with the degree of improvement correlating with the ability of oral physostigmine to increase the nocturnal secretion of cortisol. No statistically significant differences of piracetam or piracetam and lecithin, compared to placebo were noted, however, the ratio of red cell to plasma choline might be associated with treatment responsivity. The potential therapeutic efficacy of oxotremorine proved all but impossible to assess due to concomitant adverse effects, particularly dysphoria. Results with another cholinergic agonist, RS-86, will be reported. This drug appeared to be better tolerated than oxotremorine. Animals with a kianic acid induced cortical depletion of choline acetyltransferase were found to have a significant impairment in retention of a passive avoidance task, an abnormality that was readily reversible by physostigmine, oxotremorine and 4-amino-pyridine. Cysteamine, a depletor of somatostatin, also produced a comparable deficit.

Rapid thyroid hormone action in vitro in the absence of new protein synthesis.

Studies were carried out on the effect of triiodothyronine (T3) on the oxygen consumption of dispersed rat liver cells incubated for 2 hr at 37 degrees C. Thyroidectomized SD-NIH rats were kept on a low iodine diet with calcium chloride in the drinking water for 4 weeks or longer to assure hypothyroidism, verified by low serum thyroxine and T3 concentrations. Liver cells were obtained by portal vein perfusion with oxygenated collagenase-enriched Krebs-Ringer-bicarbonate buffer, after the method of Berry and Friend. Cell viability was evaluated by morphology, by trypan blue exclusion, and by biochemical parameters prior to 2-hr incubations with or without added hormone. The oxygen consumption of cell suspensions was measured with the Clark oxygen electrode after the 2-hr incubations at 37 degrees C with oxygenation of the flasks and alanine (5-10 mM) as substrate. In 31 experiments the oxygen consumption (QO2) was enhanced to 121% of control values with T3 in the medium at 3.3 nM ("physiological" level) and with an even greater effect (138% of control values) with 300-1000 nM T3 ("hyperthyroid" level). Cycloheximide at 100 microM was used to inhibit new protein synthesis by incubated hepatocytes. In 18 parallel experiments with cycloheximide blockade, no alteration of the stimulatory effect of T3 was evident. The results signify that incubated liver cells show an early response to thyroid hormone by extranuclear pathways that do not require new protein synthesis.