Effect of Aging on 24-Hour Changes in Serotonin and Dopamine Turnover, and Somatostatin and Amino Acid Content, of the Rat Hypothalamus and Pituitary Gland

The effect of aging on neurotransmitter and peptide content in the hypothalamichypophysial unit has commonly been analyzed at single time points in the 24-h cycle. Since significant changes in circadian rhythmicity occur during aging, this study aimed to examine 24-h rhythmicity in hypothalamic and pituitary serotonin (5HT) and dopamine (DA) turnover and content, and somatostatin and amino acid content in 2 months-old and 18-20 months-old rats, killed at 6 different time intervals throughout the light-dark cycle. Aged rats showed suppressed or disrupted 24-h rhythms of 5HT and DA turnover and of somatostatin, glutamate, aspartate and taurine content (anterior hypothalamus), of 5HT and DA turnover and of somatostatin, glutamate, taurine and glycine content (medial hypothalamus) and of DA turnover and amino acid content (posterior hypothalamus). Twenty-four h variations in DA, somatostatin, aspartate, GABA and glycine content of the anterior hypophysis and in all parameters tested in the neurointermediate lobe became suppressed or disrupted in aged rats. Mean values generally decreased with age, except for DA content in the anterior pituitary lobe and aspartate content in the neurointermediate lobe. Conclusions: Examination of neurotransmitter and neuropeptide content at different times of the day is needed to analyze the effects of aging in the hypothalamic-hypophysial unit.     

Age-related changes in 24-hour rhythms of norepinephrine content and serotonin turnover in rat pineal gland: effect of melatonin treatment

The 24-hour rhythms of pineal norepinephrine (NE) content and serotonin (5-HT) turnover [estimated from the ratio of 5-hydroxyindoleacetic acid (5-HIAA) to 5-HT] were studied in young (2 months) and aged (18-20 months) Wistar rats killed at 6 different time points throughout a 24-hour cycle. In the first study, significant changes dependent on the time of day were identified, with acrophases in the first half of the activity span for both parameters. Old rats showed significantly smaller mesor and amplitude of the 24-hour rhythm of pineal NE content. They also showed decreased amplitude of the pineal 5-HT turnover rhythm, in the absence of changes in mesor. In old rats, pineal 5-HT and 5-HIAA concentrations were 41-47% of those found in young rats. In a second study, young and old rats received daily intraperitoneal injections of melatonin (30 microg) or vehicle for 11 days at 19.00 h (i.e. 11 h after light on). Analyzed as a main factor in a factorial analysis of variance, both pineal NE content and 5-HT turnover decreased in old rats while pineal 5-HT turnover increased after melatonin treatment. Melatonin treatment augmented the amplitude of the 24-hour rhythm of pineal NE content by 120 and 52% in young and old rats, respectively. The amplitude of the 24-hour rhythm of pineal 5-HT turnover almost doubled after melatonin treatment in young rats and did not change in old rats. Melatonin injection did not modify the rhythm's acrophase. The results indicate that old rats had lower amplitude and lower mesor values of 24-hour variations in pineal NE content and 5-HT turnover. Melatonin treatment only partly restored pineal NE content and was devoid of activity on pineal 5-HT turnover and 5-HT and 5-HIAA concentration in old rats. Impairment of pineal melatonin synthesizing capacity and intrapineal responses to melatonin may underlie pineal aging in rats.     

Cadmium effects on 24 h changes in glutamate,aspartate, glutamine,GABA and taurine content of rat striatum

This work evaluates the possible changes in 24 h variations of striatal aspartate, glutamate, glutamine, gamma-aminobutyric acid (GABA) and taurine content after oral cadmium treatment. Male rats were submitted to cadmium exposure at two doses (25 and 50 mg/L of cadmium chloride (CdCl₂)) in the drinking water for 30 days. Control rats received cadmium-free water. After the treatment, rats were killed at six different time intervals throughout a 24 h cycle. Differential effects of cadmium on 24 h amino acid fluctuations were observed. Metal exposure modified the daily pattern of the amino acids concentration found in control animals, except for GABA and taurine with the lowest dose used. Exposure to 25 mg/L of CdCl₂ decreased mean content of aspartate, as well as GABA concentration. These results suggest that cadmium exposure affects 24 h changes of the studied amino acids concentration in the striatum, and those changes may be related to alterations in striatal function.     

Amino Acids in the Substantia Nigra of Rats with Striatal Lesions Produced by Kainic Acid

In an attempt to estimate the pool size of glutamate and other amino acids in γ-aminobutyric acid (GABA)-containing neurons, we determined the content of 12 amino acids in the bilateral substantia nigra of rats, in which unilateral striatal lesions had been made with kainic acid two weeks earlier. The assay of the amino acids (including glutamate, aspartate, glutamine, asparagine, glycine, and GABA) and ethanolamine was based on HPLC and fluorimetric detection after precolumn derivatization with o-phthaldialdehyde. The levels of all measured amino acids (except those of tyrosine, threonine, and ethanolamine) were decreased in the affected striatum, but only the levels of aspartate, taurine, and GABA were lowered in the ipsilateral substantia nigra. These results indicate that the pool size of the various amino acids in the striatonigral GABAergic pathway is small compared to their nigral content, and that in addition to GABA a significant fraction of aspartate and taurine may be confined to nerve terminals in the substantia nigra.     

Possible role of glutamate, aspartate, glutamine, GABA or taurine on cadmium toxicity on the hypothalamic pituitary axis activity in adult male rats

This work was designed to evaluate the possible changes in glutamate, aspartate, glutamine, GABA and taurine within various hypothalamic areas the striatum and prefrontal cortex after oral cadmium exposure in adult male rats, and if these changes are related to pituitary hormone secretion. The contents of glutamine, glutamate, aspartate, GABA and taurine in the median eminence, anterior, mediobasal and posterior hypothalamus, and in prefrontal cortex in adult male rats exposed to 272.7 mol l^–1 of cadmium chloride (CdCl₂) in the drinking water for one month. Cadmium diminished the content of glutamine, glutamate and aspartate in anterior hypothalamus as compared to the values found in the untreated group. Besides, there is a decrease in the content of glutamate, aspartate and taurine in the prefrontal cortex. The amino acids studied did not change in median eminence, mediobasal and posterior hypothalamus or the striatum by cadmium treatment. Plasma prolactin and LH levels decreased in rats exposed to the metal. These results suggest that (1) cadmium differentially affects amino acid content within the brain region studied and (2) the inhibitory effect of cadmium on prolactin and LH secretion may be partially explained by a decrease in the content of both glutamate and aspartate in anterior hypothalamus, but not through changes in GABA and taurine.     

Postnatal development of TRH and TSH rhythms in the rat

We previously observed that under a 12-hour light/12-hour dark schedule (lights off at 19.00 h), adult male Sprague-Dawley rats showed a circadian rhythm for serum thyroid-stimulating hormone (TSH) with a zenith near midday. In the present work, the ontogenesis of serum TSH rhythm was determined as well as pituitary TSH variations. In addition, hypothalamic and blood TRH were measured in these rats aged 15, 25, 40 and 70 days when sacrificed. As from the first age studied (15 days), a hypothalamic thyrotropin-releasing hormone (TRH) circadian rhythm was present. The mesor and the amplitude of this hypothalamic TRH rhythm increased while the rats were growing up, in contrast with the decrease observed for these parameters as far as blood TRH circadian rhythm is concerned. The time of the acrophase moved from 17.32 h in the 15-day-old rats to 13.57 h in the 70-day-old rats, being constantly in phase opposition with the blood TRH acrophase. The low amplitude pituitary TSH circadian rhythm detected in the young rat disappeared in the adult while, in contrast, the serum TSH rhythm became consistent to reach the well-characterized circadian midday peak in the 70-day-old rats.     

Toxic effects of methoxychlor in rat striatum: modifications in several neurotransmitters

Neurotoxic effects of methoxychlor (MTX) are poorly understood at present. This study was undertaken to evaluate the possible effects of MTX in norepinephrine, dopamine and amino acid contents and serotonin turnover in rat striatum. For this purpose, adult male Sprague-Dawley rats were administered 25 mg/kg/day of MTX in sesame oil or vehicle only for 30 days. The neurotransmitters of interest were measured in the striatum by HPLC. MTX decreased norepinephrine and 5-hydroxyindole acetic acid (5-HIAA) content and serotonin turnover (measured as 5-HIAA/serotonin ratio), and increased glutamate and GABA concentrations. However, the content of serotonin, aspartate, glutamine and taurine was not modified by MTX exposure. These data suggest that MTX exposure inhibits norepinephrine synthesis and serotonin metabolism. The inhibitory effect on norepinephrine could be explained, at least in part, by the increase of both GABA and glutamate contents. Further studies are needed to understand the effects of MTX on serotonin. Also a disruptive effect of MTX on the metabolisms of glutamate, aspartate, glutamine and GABA emerges.     

Pongamia pinnata influences the circadian variations of liver marker enzymes in hyperammonemic rats

In this study, the influences of Pongamia pinnata, an indigenous medicinal plant used in Ayurvedic and traditional Medicine in India, on the circadian variations of liver marker enzymes in ammonium chloride (AC) induced hyperammonemic rats were studied. Experimental rats (160 - 180 g) were divided into control, AC (daily i.p. injection of AC (100 mg kg^-1 body weight)) treated, AC + P. pinnata ethanolic leaf extract (PPEt) (300 mg kg^-1 body weight) treated and PPEt treated groups. Temporal characteristics (acrophase, amplitude and mesor) of liver marker enzymes; alkaline phosphatase (ALP), aspartate and alanine transaminases (ALT and AST) and γ-glutamyl transferase (GGT) were analyzed. Elevated liver marker enzymes (increased mesor and delayed acrophase of AST, ALT, ALP and GGT) were found in hyperammonemic rats. Administration of PPEt significantly alters these changes. Variations in acrophase, amplitude and r values were also found in control and experimental rats. The detectable circadian rhythms of hepatic marker enzymes and their alterations during AC/PPEt treatments, in the present study, deserve further investigation for the diagnosis and for the therapeutic efficacy of hyperammonemia.     

Aspartate modulates the circadian patterns of a few biochemical variables in Wistar rats

D-aspartate was used to demonstrate possible sources of excitatory input to the suprachiasmatic nucleus (SCN) in rats. Aspartate (50 mg/kg bodyweight) was orally administrated chronically for 60 days to Wistar rats and 24 h rhythmic patterns of glucose, cholesterol, total protein and aspartate transaminase (AST) were studied under light - dark (LD 12:12 h) cycle. Our results showed acrophase advances in glucose and delays in cholesterol and AST rhythms. Increased mesor and altered amplitude values were found in all rhythms; aspartate levels in the brain were found to be significantly increased in aspartate treated animals. We hypothesised that the altered biochemical rhythms in aspartate treated rats could be due to (1) modulation of neurotransmission in SCN, (2) behavioural rhythms and (3) feeding rhythms.     

Protective effect of atorvastatin on circadian regulation of liver marker enzymes and redox status in hyperlipidemic rats

Hyperlipidemia was induced in rats by administering 2% cholesterol, 20% coconut oil, and 0.125% cholic acid for 10 weeks. Atorvastatin (0.8 mg/kg b.w.) was administered orally to rats together with high-fat diet for 10 weeks. At the end of the experimental period, the circadian characteristics (acrophase, amplitude, and mesor) of liver marker enzymes (aspartate aminotransferase and alanine transferase), lipid peroxidation products (thiobarbituric acid reactive substances (TBARS), and antioxidants (superoxide dismutase, catalase, reduced glutathione, and glutathione peroxidase) were analyzed. Circadian characteristics (mesor, amplitude, and acrophase) of liver marker enzymes, TBARS, and antioxidants were altered in high-fat diet-induced rats, and the diminished amplitude along with decreased mesor levels of antioxidants were observed in high-fat diet-induced rats. Further, oral administration of atorvastatin to high-fat diet-induced rats showed the normalized mesor, amplitude, and acrophase. These findings suggest that the antihyperlipidemic potential of atorvastatin could modulate the circadian patterns of liver marker enzymes and redox status in hyperlipidemic rats.     

Temporal variations of lipid peroxidation products, antioxidants and liver marker enzymes in experimental hyperammonemic rats

Circadian variations of lipid peroxidation products: thiobarbituric acid and reactive substances (TBARS), antioxidants: reduced glutathione (GSH), superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) and liver marker enzymes such as transaminases (aspartate transaminase (AST) and alanine transaminase (ALT), alkaline phosphatase (ALP) and γ-Glutamyl transpeptidase (GGT) in circulation were analysed in control and ammonium chloride (AC) induced (100 mg/kg bodyweight) hyperammonemic rats. Elevated lipid peroxidation and liver marker enzymes (increased mesor of TBARS, AST, ALT, ALP and GGT) associated with decreased activities of antioxidants (decreased mesor of GPx, GSH, SOD and CAT) were found in hyperammonemic rats. Variations in acrophase, amplitude and r values were also found in between the control and hyperammonemic rats. These alterations clearly indicate that temporal liver marker enzymes and redox status are modulated during hyperammonemic conditions, which may also play a crucial role in disease development.     

Effect of Impact Trauma on Neurotransmitter and Nonneurotransmitter Amino Acids in Rat Spinal Cord

N-Methyl-D-aspartate (NMDA) administration exacerbates neurological dysfunction after traumatic spinal cord injury in rats, whereas NMDA antagonists improve outcome in this model. These observations suggest that release of excitatory amino acids contributes to secondary tissue damage after traumatic spinal cord injury. To further examine this hypothesis, concentrations of free amino acids were measured in spinal cord samples from anesthetized rats subjected to various degrees of impact trauma to the T9 spinal segment. Levels of excitatory and inhibitory neurotransmitter amino acids [gamma-aminobutyric acid (GABA), glutamate, aspartate, glycine, taurine] and levels of nonneurotransmitter amino acids (asparagine, glutamine, alanine, threonine, serine) were determined at 5 min, 4 h, and 24 h posttrauma. Uninjured surgical (laminectomy) control animals showed modest but significant declines in aspartate and glutamate levels, but not in other amino acids, at all time points. In injured animals, the excitatory amino acids glutamate and aspartate were significantly decreased by 5 min posttrauma, and remained depressed at 4 h and 24 h as compared with corresponding laminectomy controls. In contrast, the inhibitory amino acids, glycine, GABA, and taurine, were decreased at 5 min postinjury, had partially recovered at 4 h, and were almost fully recovered at 24 h. The nonneurotransmitter amino acids were unchanged at 5 min posttrauma and significantly increased at 4 h, with partial recovery at 24 h. At 4 h postinjury, severe trauma caused significantly greater decreases in aspartate and glutamate than did either mild or moderate injury. These findings are consistent with the postulated role of excitatory amino acids in CNS trauma.     

Locally infused taurine, GABA and homotaurine alter differently the striatal extracellular concentrations of dopamine and its metabolites in rats

Summary We studiedin vivo the effects of locally infused taurine (50, 150, and 450 mM) on the striatal dopamine and its metabolites in comparison with those of GABA and homotaurine, a GABA_A receptor agonist, in freely moving rats. The extracellular dopamine concentration was elevated maximally 2.5-, 2- and 4-fold by taurine, GABA and homotaurine, respectively. At 150 mM concentration, at which the maximum effects occurred, homotaurine increased the extracellular dopamine more than taurine or GABA. When taurine and GABA were infused simultaneously with tetrodotoxin the output of dopamine did not differ from that in the presence of tetrodotoxin alone. In comparison, tetrodotoxin did not inhibit the increase in extracellular dopamine caused by homotaurine. Furthermore, omission of calcium from the perfusion fluid inhibited the increase of extracellular dopamine caused by GABA. However, it did not block the increase of dopamine caused by taurine or homotaurine. The present study suggests that the effects of intrastriatal taurine, GABA and homotaurine on the striatal extracellular dopamine differ. Thus, these amino acids seem to affect the striatal dopaminergic neurons via more than one mechanism.     

Amino Acid Content of Nerve Endings (Synaptosomes) in Different Regions of Brain: Effects of Gabaculine and Isonicotinic Acid Hydrazide

Abstract: The amino acid content of synaptosomes was determined in six regions of rat brain, and in all regions the five predominant amino acids were glutamate, glutamine, aspartate, taurine, and GABA (γ-aminobutyrate). However, the proportions of the individual amino acids varied considerably from one region to another, the GABA content being particularly high and the taurine content low in synaptosomes from the diencephalon and mesencephalon. Administration of isonicotinic acid hydrazide to rats lowered the synaptosomal GABA level by similar amounts in all brain regions, but the administration of gabaculine resulted in a particularly long-acting elevation in GABA levels in the nerve endings of the diencephalon and mesencephalon. The possibility is raised that the high GABA levels in the nerve terminals of the diencephalon may be involved in the gabaculine-induced lowering of the body temperature of the rats. A constancy in the amount of the synaptosomal pool of "aspartate + glutamate + glutamine + GABA" was observed despite large changes in the relative amounts of the four amino acids brought about by gabaculine.     

Effect of octreotide on 24-hour growth hormone and prolactin secretory patterns in acromegalics.

Four adult patients with active acromegaly underwent studies of their 24-hour secretory pattern of hGH and Prl prior to and at the end of 3 months of treatment with the octreotide (somatostatin analog SMS 201-995) 100 micrograms s.c. every 8 h. Blood was withdrawn at 30-min intervals with the aid of a constant withdrawal pump. The best fit cosinor method was used to define the following rhythm parameters: mesor, amplitude, acrophase and periodicity. Prior to treatment, hGH secretion was increased in all patients. The mean 24-hour ranged from 9-47 ng/ml with amplitude 5.2-23 and observed maximal pulse 41-95 ng/ml. Computed rhythms were circadian in 3 patients and ultradian in 1; in 2 patients the acrophases were shifted to daytime. hPrl secretion was altered in 3 of the patients. Two had elevated mean 24-hour of 17.7 and 22.2 ng/ml, while computed rhythms showed semicircadian periodicity in 1 of them and circadian periodicity with a shift of acrophase to daytime in the other. The third patient who had normal hPrl levels, showed ultradian 8-hour periodicity. At the end of treatment there was a marked reduction in hGH secretion in 1 patient and a lesser reduction in the other 3. The rhythm was influenced by the masking effect of the drug, to yield an 8-hour period with acrophases related to injection clock time having equal amplitudes.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of various photoperiods on daily oscillations of serum corticosterone and insulin in rats.

The effect of various photoperiods on circadian rhythms of chosen parameters was investigated in laboratory rats. SPF male Wistar rats were adapted for six weeks to artificial light-dark cycles (LD 8:16, 12:12, 16:8). The light was switched on at 07.00 h in all regimens. The rats were killed at 3-hour intervals within 24 h, the serum concentration of corticosterone, insulin, glucose, food and water intake was determined. The external and computative acrophases of corticosterone varied in every photoperiod being dependent on the duration of light, the mesor values decreased in LD 16:8 in comparison with other photoperiods. The external acrophase of insulin was located 4 h after light onset in LD 8:16 and 12:12, in LD 16:8 one hour before light onset. The mesor values were approximately equal in all photoperiods. The circadian rhythms of glucose were similar in all regimens. Circadian variation of food and water consumption culminated at the same time in all regimens, the amount of food consumed in light increased with the light duration. Various photoperiods remarkably influenced circadian oscillations of corticosterone and in part food and water intake which could be considered as photoperiodic traits.     

THE FORMATION OF GLUTAMATE, ASPARTATE AND GABA IN THE RAT RETINA; GLUCOSE AND GLUTAMINE AS PRECURSORS

Abstract— The distribution of the neuroactive amino acids taurine, GABA, glycine, glutamate and aspartate, together with glutamine, have been studied in the rat retina. Peak levels of taurine were found in photoreceptor cells and of GABA and glycine in a retinal fraction enriched in amacrine cells and, synaptic terminals. In vitro , GABA formation from [³H]glutamine and [¹⁴C]glucose was also most prominent in this fraction; at 500 μ m [³H]glutamine was the better precursor.
Observations on metabolism in the photoreceptor cell layer of the tissue suggest an active turnover of glutamate, aspartate and GABA, and show that glutamine may serve as an alternative substrate to glucose here, perhaps via the GABA bypath.     

Levels of several amino acids in the cerebellar peduncles and four medullary nuclei of control and 3-acetylpyridine-treated rats

The levels of aspartate, glutamate, GABA, taurine, glycine, and alanine were determined in the inferior (ICP), middle (MCP) and superior (SCP) cerebellar peduncles, and in the inferior olive (ION), lateral reticular (LRN), lateral vestibular (LVN) and descending trigeminal (DTN) nuclei of control rats and of rats given a single intraperitoneal injection of 65 mg/kg of 3-acetylpyridine (3-AP). The content of glutamate in the MCP and SCP was 30% higher than that found for the ICP. The content of GABA was 4 to 6-fold greater in the SCP than in either the ICP or MCP. The level of taurine in the SCP was 25% lower than the value in the MCP but was not significantly lower than the value for the ICP. The levels of aspartate, glycine, and alanine were evenly distributed among the three peduncles. The contents of aspartate, glutamate, taurine, and alanine were evenly distributed among the four medullary nuclei. The level of glycine was significantly greater in the DTN than in either the LRN or LVN. The content of GABA in the ION and LVN was significantly greater than the value found for the LRN. Injection of 3-AP caused a decrease in the level of taurine of 10% in the ION and LRN, 15% in the LVN, and 25% in the DTN. No other statistically significant differences were found in the levels of the amino acids in the peduncles or medullary nuclei following 3-AP treatment. The present data do not support the idea that asparate and/or taurine are present in relatively high concentrations in the cerebellar climbing fibers.     

Chronoendocrinological studies in athletes. Circadian rhythms of HGH and IRI

The serum concentrations of HGH and IRI studied over a 24-hour span in 83 healthy athletes under normal rest conditions in a normal living environment show statistically significant rhythms, validated and quantified by cosinor analysis; for each hormone three different types of the circadian rhythm (with a peak of acrophase between: 6.30 a.m. and 1.30 p.m., 1.30 p.m. and 10.30 p.m., 10.30 p.m. and 6.30 a.m.) were observed. The IRI response to a meal load underwent cyclic variations during the day.     

The use of microdialysis for studying the regional effects of pharmacological manipulation on extracellular levels of amino acids--some methodological aspects.

The purpose of this study was to examine and validate the use of microdialysis for sampling and pharmacologically manipulating extracellular amino acids in the brain. Repeated use of microdialysis probes in acute intracerebral experiments did not significantly alter the relative recovery in vitro for the amino acids quantitated (GABA, aspartate, glutamate, glycine, taurine, and alanine). Regional differences in basal levels of some of the amino acids were detected in dialysates collected from the dorsomedial hypothalamus, striatum, and frontal cortex. The percent in vitro recoveries for the amino acids from the probes used in the three regions were not significantly different suggesting that the regional differences in basal levels of amino acids were functionally derived and not a consequence of variations in probe recovery. Perfusion with nipecotic acid, an inhibitor of GABA uptake, resulted in selective elevations in extracellular GABA in the three regions studied. Conversely, perfusion with high-potassium, a depolarizing agent, resulted in significant elevations in not only extracellular GABA but also aspartate, glutamate, and taurine. Thus, microdialysis is a method which can be employed to assess and to pharmacologically manipulate extracellular amino acids in the rat brain.