Factors influencing the inhibition of biliary glutathione efflux induced by biliary obstruction

The aim of this study was to investigate mechanisms responsible for the inhibition of biliary glutathione efflux in rats with secondary biliary cirrhosis. Rats were studied after bile duct obstruction for 28 days. The biliary secretion of reduced glutathione (GSH), oxidised glutathione (GSSG) and cysteine were completely inhibited in biliary obstructed rats. Hepatic gamma glutamyltranspeptidase (gamma-GT) activity increased significantly, but following its inhibition by acivicin administration GSH, GSSG and cysteine were still absent in bile. Biliary obstruction resulted in a significant increase of the permeability of the paracellular pathway, as shown by the higher bile/plasma ratio and hepatic clearance of [14C]sucrose. GSH and GSSG were, however, significantly lower in the carotid artery and hepatic vein of obstructed animals and the arteriovenous difference across the liver was reduced. The concentration of GSH was significantly reduced and that of GSSG increased in the liver of obstructed rats. Biliary obstruction induced an increase in the hepatic concentration of cysteine and an inhibition of both gamma glutamylcysteine synthetase and methionine adenosyl transferase activities. Dichlorofluorescein (DCF) and the GSSG/GSH ratio and thiobarbituric acid reactive substances (TBARS) concentration, markers of reactive oxygen species production and lipid peroxidation, respectively, were significantly increased. Our data indicate that increased degradation or blood reflux of glutathione do not participate in the disruption of its secretion into bile and support the view that impairment of glutathione synthesis and oxidative stress could contribute to the decline in biliary glutathione output.     

Intrabiliary glutathione hydrolysis. A source of glutamate in bile

High concentrations of glutathione (GSH) and two of its constituent amino acids, glutamate and glycine, are normally found in rat bile. To examine the role of intrabiliary GSH hydrolysis as a source of these amino acids, as well as of cystine in bile, the biliary excretion of GSH and free amino acids was measured in normal male Sprague-Dawley rats; in animals given either phenol 3,6-dibromphthalein disulfonate or diethyl maleate, inhibitors of GSH secretion into bile; and after a retrograde intrabiliary infusion of (alpha S, 5S)-alpha-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid (AT-125), an irreversible inhibitor of gamma-glutamyl transferase activity. Total concentration of amino acids in normal rat bile ranged from 4 to 7 mM and was more than double the concentration in plasma (2-3 mM). Although most amino acids were detected in bile, glutamate and glycine were the most prevalent (1.2 and 1.0 mM, respectively), followed by the branched chain amino acids valine and leucine. The administration of phenol 3,6-dibromphthalein disulfonate (180 mumol/kg, intravenous), or of diethyl maleate (1 mmol/kg, intraperitoneal), resulted in a marked decrease in the biliary excretion of GSH, as well as a decrease in the excretion of glutamate, cystine, and glycine; however, the effects of these agents were not specific for the amino acid constituents of GSH. Following retrograde intrabiliary infusion of AT-125 (10 mumol/kg), there was an immediate and sustained doubling in the rate of biliary excretion of both GSH and glutathione disulfide and a marked decrease in the rate of excretion of glutamate. Varying the dose of AT-125 (0-20 mumol/kg) resulted in an inverse linear relation between hepatic gamma-glutamyl transferase activity and the biliary excretion of intact GSH. These findings suggest that most, if not all, of the free glutamate in excreted bile is formed from the intrabiliary hydrolysis of GSH. Prior to hydrolysis within the biliary tree, substantial concentrations of GSH must be transported from liver cells into bile; minimal canalicular concentrations of this tripeptide are estimated at 5 mM.     

Age-related changes of NA(+), K(+) - ATPase, Ca(+2) - ATPase and Mg(+2) - ATPase activities in rat brain synaptosomes.

Cerebral metabolism of glucose, one of the determinants of tissue ATP level, is crucial for central nervous system function. The activity of P-type pumps, namely Na(+), K(+) - ATPase, Ca(+2) - ATPase and Mg (+2) - ATPase were examined in brain synaptosomes of 5 - day, 3 - month and 18 - month - old rats to determine if changes in enzyme activity related to aging are potentially associated with alterations in glucose homeostasis. Activities of all the ATPases studied in isolated brain synaptosomes were expressed in micromol of Pi liberated from ATP by 1 mg of synaptosome protein during one hour. Serum glucose concentration was measured by the glucose oxidase method and insulin level was estimated by the RIA. Our results demonstrate that 18 - month - old rats are characterized by hyperglycemia and hyperinsulinemia. Their serum glucose concentration was significantly increased approx. 62.3% and 135.8 % as compared to 3 - month - old rats and 5 - day, newborn rats, respectively. An enormous increase in serum insulin concentration in the old, hyperglycemic rats was observed concomitantly. As a result of these changes the insulin - to - glucose ratio in the old rats was greatly increased approx. (270% and 230%) compared to young, mature and newborn rats. Hyperglycemia and hyperinsulinemia occurring in the old rats, had a different impact on activities of the ATPases tested. Our results have revealed that Na(+), K(+) - ATPase activity remains almost unchanged with age, the activity of Ca(+2) - ATPase decreases, whereas that of Mg(+2) - ATPase increases significantly in old, insulin resistant rats. In conclusion it seems that changes in activity of different P - type pumps may differ with aging and that adaptation of specific ATPases to internal environment alterations is not identical.     

Role of S-adenosylmethionine on the hepatobiliary homeostasis of glutathione during cyclosporine A treatment

The effects of cyclosporine A (CyA) treatment on the hepatic content and biliary output of reduced (GSH) and oxidized (GSSG) glutathione and lipid peroxidation in the liver, and the ability of S-adenosylmethionine (SAMe) to antagonize the CyA-induced alterations were studied in male Wistar rats. To evaluate the efficacy of SAMe, three CyA and SAMe protocols were used: cotreatment with SAMe plus CyA, pretreatment with SAMe before starting cotreatment, and post-treatment with SAMe after beginning treatment with CyA alone. CyA treatment for one and four weeks depleted liver GSH, decreased the GSH/GSSG ratio and significantly reduced GSH and GSSG biliary concentrations and secretion rates. Additionally, long-term treatment enhanced lipid peroxidation. By contrast, when the rats were treated with CyA plus SAMe using any of the administration protocols, SAMe was seen to be efficient in antagonizing the GSH hepatic depletion, the changes in hepatic GSH/GSSG ratio and the increase induced by CyA in lipid peroxidation. Furthermore, SAMe also abolished the effects of CyA on the biliary secretion rates of GSH and GSSG. The efficacy of SAMe was similar, regardless of the administration protocols used. In conclusion, our results clearly demonstrate that SAMe is good for preventing, antagonizing and reversing the CyA-induced alterations in the hepatobiliary homeostasis of glutathione.     

Differential effects of mild therapeutic exercise during a period of inactivity on power generation in soleus type I single fibers with age

The purpose of this study was to investigate the effects of mild therapeutic exercise (treadmill) in preventing the inactivity-induced alterations in contractile properties (e.g., power, force, and velocity) of type I soleus single fibers in three different age groups. Young adult (5- to 12-mo-old), middle-aged (24- to 31-mo-old), and old (32- to 40-mo-old) F344BNF1 rats were randomly assigned to three experimental groups: weight-bearing control (CON), non-weight bearing (NWB), and NWB with exercise (NWBX). NWB rats were hindlimb suspended for 2 wk, representing inactivity. The NWBX rats were hindlimb suspended for 2 wk and received therapeutic exercise on a treadmill four times a day for 15 min each. Peak power and isometric maximal force were reduced following hindlimb suspension (HS) in all three age groups. HS decreased fiber diameter in young adult and old rats (-21 and -12%, respectively). Specific tension (isometric maximal force/cross-sectional area) was significantly reduced in both the middle-aged (-36%) and old (-23%) rats. The effects of the mild therapeutic exercise program on fiber diameter and contractile properties were age specific. Mild treadmill therapeutic exercise attenuated the HS-induced reduction in fiber diameter (+17%, 93% level of CON group) and peak power (μN·fiber length·s(-1)) (+46%, 63% level of CON group) in young adult rats. In the middle-aged animals, this exercise protocol improved peak power (+60%, 100% level of CON group) and normalized power (kN·m(-2)·fiber length·s(-1)) (+45%, 108% level of CON group). Interestingly, treadmill exercise resulted in a further reduction in shortening velocity (-42%, 67% level of CON group) and specific tension (-29%, 55% level of CON group) in the old animals. These results suggest that mild treadmill exercise is beneficial in attenuating and preventing inactivity-induced decline in peak power of type I soleus single fibers in young adult and middle-aged animals, respectively. However, this exercise program does not prevent the HS-induced decline in muscle function in the old animals.     

Decreased response with age of the cardiac catecholamine sensitive adenylate cyclase system

The cardiac β-adrenergic coupled adenylate cyclase system was examined in young and old male Wistar rats. The concentration of binding sites for (?) ³H-DHA in membranes prepared from cardiac ventricles was 21.1 ± 2.78 (SD) fmoles/mg protein in 3–4 month old rats (young rats) and 31.2 ± 2.20 fmoles/mg protein in 24 month old rats (old rats). The dissociation constant, K_D was 4.3 ± 1.8 nM and 6.7 ± 1.7 nM for young and old rats, respectively. Various compounds were used to study the characteristics of activation of adenylate cyclase in homogenates from cardiac ventricles. Basal adenylate cyclase was reduced 30% in old animals compared to young (6.1 pmoles/min/mg protein in 24 month vs. 8.6 pmoles/min/mg protein in 3–4 month). (?)Isoproterenol (10^?5M) alone stimulated adenylate cyclase greater than two-fold in young rats (10.6 pmoles/min/mg protein above basal) and this stimulation was 34% lower in old animals. GppNHp (100 μM), fluoride (10 mM), and forskolin (100 μM) activation of adenylate cyclase above basal was reduced 38, 37, and 34%, respectively, in the old animals. No significant changes between the two groups were noted in the apparent affinity of GppNHp either alone or in the presence of (?)isoproterenol nor in the affinities of catecholamine agonists for activation of cyclase. These results suggest a reduction in the amount of functional regulatory protein or possibly cyclase in 24 month old rat ventricular tissue compared to 3–4 month old tissue. However, this data does not rule out the possibility of altered molecular interactions of a full complement of regulatory protein (s) with β-adrenergic receptor and/or catalytic adenylate cyclase.     

The effects of reserpine and haloperidol on tyrosine hydroxylase activity in the brains of aged rats

Many neurotransmitter systems appear to be altered with aging. The effects of aging on the regulation of tyrosine hydroxylase, the rate-limiting enzyme in the synthesis of catecholamines in the brain has been examined. The endogenous basal activity of tyrosine hydroxylase was lower in the hypothalamus of 24 month old Fisher 344 rats than in the hypothalamus of 3 month old or 6 month old animals. There was no difference in the basal activity of tyrosine hydroxylase in the locus ceruleus, frontal cortex, hippocampus, substantia nigra, or the striatum of rats of ages 3 months, 6 months and 24 months. Tyrosine hydroxylase activity was increased in the striatum of 3 month old (60%) and 6 month old (28%) rats after treatment with haloperidol or reserpine, whereas no change in enzyme activity followed administration of these drugs to 24 month old animals. In conclusion, increases in tyrosine hydroxylase activity in the brain that normally occur in the striatum of 3 month old rats after haloperidol or reserpine treatment are significantly decreased in 6 month old rats and not apparent in 24 month old rats.     

Effects of aging on the susceptibility to the toxic effects of cyclosporin A in rats. Changes in liver glutathione and antioxidant enzymes

Free radicals are involved in aging and cyclosporin A-induced toxicity. The age-related changes in the liver oxidative status of glutathione, lipid peroxidation, and the activity of the enzymatic antioxidant defense system, as well as the influence of aging on the susceptibility to the hepatotoxic effects of cyclosporin (CyA) were investigated in rats of different ages (1, 2, 4, and 24 months). The hepatic content of reduced glutathione (GSH) increased with aging, peaked at 4 months, and decreased in senescent rats. By contrast, glutathione disulfide (GSSG) and thiobarbituric acid-reactive substances (TBARS) concentrations and superoxide dismutase, catalase, and glutathione peroxidase activities were higher in the oldest than in the youngest rats. CyA treatment, besides inducing the well-known cholestatic syndrome, increased liver GSSG and TBARS contents and the GSSG/GSH molar ratio, and altered the nonenzymatic and enzymatic antioxidant defense systems. The CyA-induced cholestasis and hepatic depletion of GSH, and the increases in the GSSG/GSH ratio, and in GSSG and TBARS concentrations were higher in the older than the mature rats. Moreover, superoxide dismutase and catalase activities were found to be significantly decreased only in treated senescent rats. The higher CyA-induced oxidative stress, lipoperoxidation, and decreases in the antioxidant defense systems in the aged animals render them more susceptible to the hepatotoxic effects of cyclosporin.     

Acetaminophen hepatotoxicity in vivo is not accompanied by oxidant stress

Hepatotoxic doses of acetaminophen in Fischer 344 rats did not increase biliary efflux of oxidized glutathione. Pretreatment of the animals with bis(2-chloroethyl)-N-nitrosourea inhibited hepatic glutathione reductase by 73 percent but did not potentiate the hepatotoxicity of acetaminophen and did not produce an increase in biliary efflux of oxidized glutathione in response to acetaminophen. Hepatic protein thiol content was not depleted by acetaminophen. A proposed role for oxidant stress mechanisms mediated either by reactive oxygen species or by the direct oxidant action of a reactive metabolite in acetaminophen-induced hepatotoxicity is unsubstantiated and unlikely.     

Effect of diethyl ether on the biliary excretion of acetaminophen

The biliary and renal excretion of acetaminophen and its metabolites over 8 hr was determined in rats exposed to diethyl ether by inhalation for 1 hr. Additional rats were anesthetized with urethane (1 g/kg ip) while control animals were conscious throughout the experiment (surgery was performed under hexobarbital narcosis: 150 mg/kg ip; 30-min duration). The concentration of UDP-glucuronic acid was decreased 80% in livers from ether-anesthetized rats but was not reduced in urethane-treated animals when compared to that in control rats. The concentration of reduced glutathione was not affected by either urethane or diethyl ether. Basal bile flow was not altered by the anesthetic agents. Bile flow rate after acetaminophen injection (100 mg/kg iv) was increased slightly over basal levels for 2 hr in hexobarbital-treated control rats, was unaltered in urethane-anesthetized animals, and was decreased throughout the 8-hr experiment in rats exposed to diethyl ether for 1 hr. In control and urethane-anesthetized animals, approximately 30-35% of the total acetaminophen dose (100 mg/kg iv) was excreted into bile in 8 hr, while only 16% was excreted in rats anesthetized with diethyl ether. Urinary elimination (60-70% of the dose) was not altered by exposure to ether. Separation of metabolites by reverse-phase high-pressure liquid chromatography showed that ether decreased the biliary elimination of unchanged acetaminophen and its glucuronide, sulfate, and glutathione conjugates by 47, 40, 49, and 73%, respectively, as compared to control rats. Excretion of unchanged acetaminophen and the glutathione conjugate into bile was depressed in urethane-anesthetized animals by 45 and 66%, respectively, whereas elimination of the glucuronide and sulfate conjugates was increased by 27 and 50%, respectively. These results indicate that biliary excretion is influenced by the anesthetic agent and that diethyl ether depresses conjugation with sulfate and glutathione as well as glucuronic acid.     

Thromboxane receptor blockade improves cyclosporine nephrotoxicity in rats

Cyclosporine A (CyA) nephrotoxicity is associated with impaired renal hemodynamic function and increased production of the vasoconstrictor eicosanoid thromboxane A2 (TxA2). In CyA toxic rats, renal dysfunction can be partially reversed by inhibitors of thromboxane synthase. However, interpretation of these results is complicated since inhibition of thromboxane synthase may cause accumulation of prostaglandin endoperoxides that can act as partial agonists at the TxA2 receptor and may blunt the efficacy of treatment. Furthermore, these endoperoxides may be used as substrate for production of vasodilator prostaglandins causing beneficial effects on hemodynamics which are independent of thromboxane inhibition. To more specifically examine the role of TxA2 in CyA toxicity, we investigated the effects of the thromboxane receptor antagonist GR32191 on renal hemodynamics in a rat model of CyA nephrotoxicity. In this model, administration of CyA resulted in a significant decrease in glomerular filtration rate (GFR) (2.85 +/- 0.26 [CyA] vs 6.82 +/- 0.96 ml/min/kg [vehicle]; p less than 0.0005) and renal blood flow (RBF) (21.65 +/- 2.31 [CyA] vs 31.87 +/- 3.60 ml/min/kg [vehicle]; p less than 0.025). Renal vascular resistance (RVR) was significantly higher in rats given CyA compared to animals treated with CyA vehicle (5.32 +/- 0.55 [CyA] vs. 3.54 +/- 0.24 mm Hg/min/ml/kg [vehicle]; p less than 0.05). These renal hemodynamic alterations were associated with a significant increase in urinary excretion of unmetabolized, "native" thromboxane B2 (TxB2) (103 +/- 18 [CyA] vs 60 +/- 16 pg/hour [vehicle]; p less than 0.05). Only minimal histomorphologic changes were apparent by light microscopic examination of kidneys from both CyA and vehicle treated animals. However, with immunoperoxidase staining, a significantly greater number of cells expressing the rat common leukocyte antigen was found in the renal interstitium of rats given CyA. There was no detectable increase in monocytes/macrophages in the kidneys of CyA toxic animals. In rats treated with CyA, intraarterial infusion of GR32191 at maximally tolerated doses significantly increased GFR and RBF, and decreased RVR. Although both RBF and RVR were restored to levels not different from controls, GFR remained significantly reduced following administration of GR32191. These data suggest that the potent vasoconstrictor TxA2 plays an important role in mediating renal dysfunction in CyA nephrotoxicity. However, other factors may be important in producing nephrotoxicity associated with CyA.     

Depression of biliary glutathione excretion by chronic ethanol feeding in the rat

The effects of chronic alcohol feeding on biliary glutathione excretion were studied in rats pair fed diets containing either ethanol (36% of total energy) or isocaloric carbohydrate for 4-6 weeks. An exteriorized biliary-duodenal fistula was established and total glutathione (GSH) and oxidized glutathione (GSSG) were measured. A significant decrease was observed in rats fed alcohol chronically compared to their pair fed controls in the biliary excretion of GSH (55.7 +/- 37.0 vs 243.1 +/- 29.0 micrograms/ml bile, p less than 0.025) as well as biliary GSSG (12.5 +/- 5.0 vs 49.9 +/- 8.0 micrograms/ml bile, p less than 0.05) and in bile flow (23.1 +/- 1.6 vs 29.2 +/- 1.3 micrograms/min, p less than 0.05). An acute dose of ethanol tended to exaggerate the decrease on biliary GSH and GSSG in the two groups of animals. The depression in biliary GSH could not be attributed to decreased GSH synthesis since S35-L-methionine incorporation into hepatic and biliary GSH was unchanged or even increased after chronic ethanol feeding.     

Age related toxic effects of endosulfan on certain enzymes of rat erythrocytes

The effect of oral administration of endosulfan (12.5 mg/kg body weight), daily for 4 days was investigated on erythrocytes of female rats of 4 different age groups i.e. 15, 30, 70 and 365 days old. Erythrocyte membrane Na+, K(+)-ATPase and Mg2(+)-ATPase activities were significantly inhibited in all the age groups of rats. However, percent inhibition was maximum in the youngest animals. A significant decrease in the activity of erythrocyte glutathione reductase was observed in 30 and 70 days old rats whereas a significant increase in the activity of glucose-6-phosphate dehydrogenase (G-6-PD) was observed in these groups. The increase in G-6-PD activity may be a physiological response to compensate for decrease in the reduced glutathione level which results from decrease in the activity of glutathione reductase.     

Antioxidant and antiplatelet effects of the alpha-tocopherol-aspirin combination in type 1-like diabetic rats

We analyze the effect of the combination of acetylsalicylic acid (2 mg/kg/day p.o.) and alpha-tocopherol (25 mg/kg/day p.o.) in a type-1-like experimental model of diabetes mellitus on platelet factors, endothelial antithrombotic factors and tissue oxidative stress. In diabetic rats, the combination of drugs had a greater inhibitory effect on platelet aggregation than in untreated control animals with diabetes (88.87%). The combination of drugs had little effect on the inhibition of thromboxane production (-90.81%) in comparison to acetylsalicylic acid alone (-84.66%), potentiated prostacyclin production (+162%) in comparison to alpha-tocopherol alone (+30.55%), and potentiated nitric oxide production (+241%) in comparison to either drug alone (acetylsalicylic acid +125%, alpha-tocopherol +142%). The combination of the two drugs improved the thromboxane/prostacyclin balance (0.145+/-0.009) in comparison to untreated diabetic animals (4.221+/-0.264) and in untreated healthy animals (0.651+/-0.045). It did not potentiate the antioxidant effect of either drug alone, but did increase tissue concentrations of reduced glutathione, especially in vascular tissue (+90.09% in comparison to untreated animals). In conclusion, in the experimental model of diabetes tested here, the combination of acetylsalicylic acid and alpha-tocopherol led to beneficial changes that can help protect tissues from thrombotic and ischemic phenomena.     

The role of old age in the effects of glucose on insulin secretion, pentosephosphate shunt activity, pyridine nucleotides and glutathione of rat pancreatic islets

In pancreatic islets of adult (three month) and old (24 month) rats the effect of glucose on glucose oxidation, pyridine nucleotides, glutathione and insulin secretion was studied. DNA content was similar in both groups of animals; however, islets of old rats exhibited 30% less insulin content. While glucose-induced (16.7 mM) insulin secretion in islets of old rats was approximately 50% less than in islets of adults, no significant difference was observed in the insulin releasing effect of theophylline (1 mM). Although islet production of 14CO2 in the presence of 16.7 mM glucose increased equally in both groups, elevation of glucose failed to increase the percentage of total glucose oxidation via the pentose phosphate shunt in islets of old rats. Elevation of glucose increased the NADPH/NADP and the NADH/NAD ratio in both groups of islets in a similar manner. The effect of glucose on the GSH/GSSG ratio revealed a dose-related increase in the islets of adult rats, whereas islets of old rats did not respond to elevation of glucose. Our data seem to indicate that the lower secretory response of islets of old rats is related to the failure of glucose to increase the GSH/GSSG ratio. In contrast the insulin release induced by theophylline does not appear to depend on islet thiols.     

Dehydroepiandrosterone improves hepatic antioxidant reserve and stimulates Akt signaling in young and old rats

This study examined, in the liver of young and old (3- and 24-month-old, respectively) healthy Wistar rats, the in vivo effect of dehydroepiandrosterone (DHEA) (10mg/kg body weight) administered subcutaneously for 5 weeks. Reduced (GSH) and oxidized (GSSG) glutathione levels, glucose-6-phosphate dehydrogenase (G6PDH), glutathione-S-transferase (GST), glutathione peroxidase (GPx) and catalase (CAT) activities, hydrogen peroxide concentration, GST and p-Akt/Akt immunocontent ratio were assessed in hepatic tissue. DHEA treatment significantly increased total glutathione content (17%) and GSH (22%) in 3- and 24-month-old treated groups when compared to control groups. The aging factor increased G6PDH (51%) and GPx (22%) activities as well as the hydrogen peroxide concentration (33%), independently of treatment. DHEA treatment increased p-Akt (54%) and p-Akt/Akt ratio (36%) immunocontents in both treated groups. Increased serum levels of alanine aminotransferase (ALT) in aged rats were reduced by DHEA treatment (34%).     

Effects of age on morphology,protein synthesis and secretagogoue-evoked secretory responses in the rat lacrimal gland

This study investigated changes in the morphology and protein synthesis and protein and peroxidase secretion due to peptidergic and aminergic stimulation from rat lacrimal gland acinar cells of 3–5, 9, 12, 20 and 24 month old rats. There was a marked reduction in the presence of Golgi apparatus in the acinar cells of glands from the 24 month old rats coupled to dilatation and degeneration of rough endoplasmic reticulum, when compared to that in the acinar cells of glands from 3–5 and 12 month old rats. Following incorporation of tritiated leucine for 360 min (6 h), the amount of newly synthesised protein in acinar cells of the 12 month old rats was significantly (p < 0.01) higher than that in the acinar cells of 3–5 month old animals. However, at 20 months the amount of newly synthesised protein in these acinar cells was significantly (p < 0.01) reduced to less than that in acinar cells of both the 3–5 and 12 month old animals. Immunohistochemical and immunofluorescence studies identified the presence of substance P (SP), vasoactive intestinal peptide (VIP), histamine and 5-hydroxytryptamine (5-HT) in the lacrimal glands of 3–5 month old rats. Stimulation by either SP, VIP, histamine or 5-HT resulted in significant increases in total protein output and peroxidase release from acinar cells of the 3–5 month old rats. However, all responses to the secretagogues were reduced with ageing from 3–5 to 24 months of age. The results indicate that ageing is associated with alteration in the ability of acinar cells to synthesise and secrete proteins.     

In Vivo Release of Endogenous Amino Acids from the Rat Striatum: Further Evidence for a Role of Glutamate and Aspartate in Corticostriatal Neurotransmission

By means of the push-pull cannula method, the outflow of endogenous amino acids was studied in the striatum of halothane-anesthetized rats. Addition of K+ ions (30 mM for 4 min) to the superfusion fluid increased the release of aspartate (+116%), glutamate (+217%), taurine (+109%), and gamma-aminobutyric acid (GABA) (+429%) whereas a prolonged decrease in the outflow of glutamine (-28%) and a delayed reduction in the efflux of tyrosine (-25%) were observed. In the absence of Ca2+, the K+-induced release of aspartate, glutamate, and GABA was blocked whereas the K+-induced release of taurine was still present. Under these conditions, the decrease in glutamine efflux was reduced and that of tyrosine was abolished. Local application of tetrodotoxin (5 microM) decreased only the outflow of glutamate (-25%). One week following lesion of the ipsilateral sensorimotor cortex the spontaneous outflow of glutamine and of tyrosine was enhanced. Despite the lack of change in their spontaneous outflow, the K+-evoked release of aspartate and glutamate was less pronounced in lesioned than in control animals, whereas the K+-evoked changes in GABA and glutamine efflux were not modified. Our data indicate that the push-pull cannula method is a reliable approach for the study of the in vivo release of endogenous amino acids. In addition, they provide further evidence for a role for glutamate and aspartate as neurotransmitters of corticostriatal neurons.     

Monoamine Transporter Gene Expression in the Central Nervous System in Diabetes Mellitus

Abstract: Diabetic animals exhibit altered neurotransmission in brain monoaminergic systems. By means of in situ hybridization, we have investigated the expression of dopamine, noradrenaline, and serotonin transporter (DA-T, NA-T, and 5-HT-T, respectively) mRNAs in the brains of alloxan- and streptozotocin-diabetic rats. The expression of DA-T mRNA is decreased in 1- (−11%) and 4-(−17%) week alloxan-diabetic and 4- (−9%) and 8-(−20%) week streptozotocin-diabetic rats in the ventral medial bundle. The expression of NA-T mRNA is decreased in the locus coeruleus of 8- (−26%) week streptozotocin-diabetic rats, in the noradrenergic A1 cell group of 4-(−27%) week alloxan- and 8- (−25%) week streptozotocin-diabetic rats, and in the noradrenergic A2 cell group of 1- (−21%) and 4- (−28%) week alloxan-diabetic and 4- (−27%) and 8- (−25%) week streptozotocin-diabetic animals. The expression of 5-HT-T mRNA in the dorsal raphe nucleus is increased in 1- (+14%) and 4- (+44%) week alloxan- and 4- (+28%) and 8-(+44%) week streptozotocin-diabetic rats. The expression of each of the three monoamine transporter genes may be differentially regulated in diabetes and dependent on the duration of diabetes. Altered monoamine transporter gene expression may possibly contribute to the observed dysfunctions in brain monoamine transmission in chronic diabetes.