Diurnal variations in the catecholamine content in rat tissues. Effects of exogenous noradrenaline

Endogenous noradrenaline (NA) content of the interscapular brown adipose tissue (IBAT), and of the hypothalamus, as well as adrenaline (A) and NA content of the adrenals were measured in the morning and in the evening in rats treated with saline (controls) or with NA (1.6 mg kg-1 pp). NA content in IBAT and in hypothalamus of control animals varies diurnally. NA content in IBAT was significantly higher in the evening than in the morning. In the hypothalamus, these variations were different of those found in the IBAT. Exogenous NA applied both in the morning and in the evening increases significantly NA content in IBAT both in the morning and in the evening, but more in the evening. Unlike IBAT, NA content of the hypothalamus after NA administration does not change essentially. In the adrenal gland of control rats, A content only is changed during the day, being markedly lower in the evening. After NA injection both A and NA are not changed significantly.     

Sympatho-adrenal activity and metabolic responses in fasted rats--the role of interscapular brown adipose tissue

1. Sympatho-adrenal (SA) and metabolic responses to fasting were studied in sham-operated (SHAM) rats and in those with interscapular brown adipose tissue (IBAT) removed. 2. Fasting significantly increased adrenaline (A) excretion and serum free fatty acids (FFA), but decreased noradrenaline (NA) excretion and blood glucose level in SHAM rats. 3. IBAT removal did not change metabolic responses while it markedly altered the SA activity. Fasting in animals void of IBAT potentiated the activity of adrenal medulla, inhibited the FFA rise and prevented glucose reduction, which is normally observed in SHAM-fasted rats. 4. Results suggest the significance of IBAT in the regulation of the blood level of energy substrates in fasted rats and in maintaining the basal level of NA excretion.     

Adrenal catecholamine synthesis rate changes induced by combined thermal and immobilization stress in fed and 24 hour fasted rats

The combined stress of acute immobilization (IM) at high and low ambient temperature has been used to determine its influence on adrenal catecholamine (CA) content assassed histofluorimetrically in fed and 24 hour fasted rats. The general course of changes obtained after the arrangement of adrenal strips deriving from the adrenals of rats exposed to cold and IM stress (CIMS) at +10 degrees C to -25 degrees C during the different time fragments presented the adrenal CA depletion events followed sometimes by the adrenal CA content increase after the longer stress exposure or/and stronger CIMS and WIMS conditions. It was found that this depletion-stimulated increase of adrenal Ca synthesis rate had been accelerated in 24 h fasted rats compared to satiated ones exposed to the same stress conditions, especially after the CIMS exposure. Moreover the survival time duration at first lethal temperature (-5 degrees C and +45 degrees C) was significantly higher in fasted rats. The possible hypothalamic regulation of adrenal CA synthesis rate accordingly to the actual metabolism needs and beta-adrenoceptor sensitivity changes depending on satiety state have been discussed and the necessity of further investigations concerning the specificity of stress-induced metabolism changes in 24 h starved rats has been suggested.     

Influence of amygdala catecholamines on ovarian and adrenal medullary secretion.

The amygdaloid complex participates in the modulation of endocrine functions, and contains measurable amounts of noradrenaline (NA) and dopamine (DA). This study examined the contribution of the amygdaloid catecholaminergic systems to the regulation of the adrenal medulla and the ovary. To accomplish this the neurotoxin 6-hydroxydopamine (6-OHDA) was bilaterally injected into the basolateral nucleus of the amygdala (ABL) in cycling rats. The contents of NA and DA in right and left amygdala decreased significantly in lesioned animals with respect to sham lesioned animals, but hypothalamic levels were not different between groups. Administration of 6-OHDA to rats increased the NA, DA and adrenaline (A) contents of the adrenals compared to vehicle treated rats. In addition, lesioned animals showed a significant increase of NA and DA contents in the ovary, although A levels did not differ between groups. Serum oestradiol (O) concentrations were significantly lower in lesioned animals than in controls. These data suggest that the amygdaloid catecholaminergic systems exert an inhibitory effect on catecholamine content of the adrenals and the ovary, and influence the ovarian oestradiol secretion mechanism.     

Effects of fasting on aldosterone secretion in ovariectomized rats

The present study was designed to assess the effect of fasting on aldosterone secretion in ovariectomized (Ovx) rats. Ovx rats were divided into fed (allowed access to food ad libitum) and fasted (deprived of food for 24 hours) groups. The trunk blood of fed and fasted rats was collected after decapitation. In the in vitro study, adrenal zona glomerulosa (ZG) cells from fed or fasted rats were incubated with angiotensin II (Ang II, 10(-6) M), adrenocorticotropic hormone (ACTH, 10(-9) M), or forskolin (an activator of adenylyl cyclase, 10(-6) M) at 37 degrees C for 30 min. The levels of aldosterone in medium and plasma extracts were measured by radioimmunoassay. Results showed that the levels of plasma aldosterone in fasted rats were lower than those in fed rats. There were no significant differences in basal and Ang II-stimulated aldosterone secretion between fed and fasted groups. The increment of aldosterone induced by ACTH in fasted group was significantly less than that in fed group. Administration of forskolin led to a significant increase in aldosterone secretion in both fed and fasted groups. Fasted group had a decreased aldosterone secretion in response to forskolin as compared with fed group. In summary, these results suggest that fasting decreases aldosterone secretion in Ovx rats through a mechanism in part involving a reduction of aldosterone production in response to ACTH, a decreased activity of adenylyl cyclase, and/or an inhibition of post-cAMP pathway in ZG cells.     

24 hour fasting and adrenoreceptor blocking agent influence on adrenal catecholamine synthesis rate changes induced by combined thermal and immobilization stress in rats.

The catecholamine (CA) depletion degree in rat adrenal medulla, the survival time (ST) and rectal temperature changes induced by combined thermal and immobilization stress were examined with the aim to prove tha alpha- and beta-adrenoreceptor sensitivity decrease implication in 24 hour fasting-induced changes of the above mentioned phenomena. The significant ST increase in strong stressful situations and the adrenal CA turnover augmentation in fed and fasted-propranolol or dihydroergotamine pretreated rats compared to untreated ones provided evidence that alpha- and beta adrenoceptor blockade causes adrenal CA turnover increase and ST prolongation similar to effects observed in our previous experiments. Thus the implication of fasting induced adrenoreceptor downregulation in adrenal CA turnover augmentation and ST prolongation of fasted rats in strong stressful conditions was suggested and the possible mechanisms of these phenomena have been discussed.     

Catecholamine and thyroid hormone influence on brown fat Na+, K+-ATPase activity and thermogenesis in the rat

Resting oxygen consumption (VO2) before and after injection of noradrenaline (NA), and plasma triiodothyronine levels were elevated in hyperthyroid and hyperphagic cafeteria fed rats, but were reduced in 4d-fasted and hypothyroid animals compared to controls. Refeeding fasted rats with a single carbohydrate meal caused all of these parameters to increase towards control levels. In vivo turnover, and in vitro release of NA brown adipose tissue (BAT) was elevated in cafeteria fed rats but remained unaltered in other groups and levels and uptake of NA in BAT were similar for all rats. Basal and NA stimulated Na+,K+-ATPase activity in BAT was increased in cafeteria and hyperthyroid rats and reduced in fasted and hypothyroid animals compared to control and refed groups. A highly significant correlation (r = 0.977), (P less than 0.001), found between the in vitro activity of this enzyme and resting VO2 in all rats, indicates that BAT Na+,K+-ATPase may be involved in the thermogenic responses to diet, catecholamines and thyroid hormones.     

Species differences in the distribution of adrenal free and sulfoconjugated catecholamines

Summary We determined free and sulfoconjugated catecholamines (CA) in adrenals of several species by reverse-phase high performance liquid chromatography (HPLC) with electrochemical detection. The two main adrenal CA, free epinephrine (E) and norepinephrine (NE) from eight species (guinea-pig, rat, dog, mice, bovine, cat, green-monkey and human) were considerably different both in the total amount as well as their relative proportions. Free dopamine (DA) content also differed from species to species but this CA was present in a relatively constant proportion, representing about 1% of the total free CA. Phenolsulfotransferase (PST) activity was present in all of the adrenals. Sulfoconjugated CA, however, were only selectively present: E and NE sulfate were entirely absent but in most of these species DA sulfate was detected in a proportion corresponding to 1–10% of the total (free + sulfoconjugated) DA. The adrenal DA sulfate concentrations did not parallel the adrenal PST activity, indicating that this enzyme can not be considered to be an index of the CA sulfates present in this organ.Abbreviations CA catecholamines - DA dopamine - DHBA dihydroxybenzylamine - E epinephrine - HPLC high performance liquid chromatography - NE norepinephrine - PST phenolsulfotransferase     

Interaction between exercise training and cold acclimation in rats

Five groups of 10 rats were used. Group A included sedentary rats kept at 24 degrees C, group B exercised-trained rats and group C rats exposed at -15 degrees C for 2 h every day and kept at 24 degrees C for the remaining time. These 3 groups were kept on this regimen for 10 weeks. In addition group D was acclimated to cold (2 h.d-1 at -15 degrees C) for 6 weeks and subsequently deacclimated at 24 degrees C for 4 weeks. Group E was also acclimated to cold for 6 weeks and during the deacclimation, at 24 degrees C period which lasted 4 weeks, the animals were exercised 2 h per day. Following the 10 week experimental period all animals were sacrificed and DNA and protein content of the IBAT as well as its total mass were measured. The results show significant increases in the cold adapted group. Exercise training which had no effect on brown adipose tissue IBAT at room temperature, caused an accelerated reduction in weight, DNA and protein content of the BAT in rats previously acclimated to cold. In spite of this, the thermogenic response to noradrenaline was significantly enhanced in the group which exercised during the deacclimation period. It is suggested that tissues other than IBAT may explain this enhanced heat production capacity.     

Shivering and nonshivering thermogenesis in exercised cold-deacclimated rats

Norepinephrine (NE)-induced increase in oxygen consumption (VO2) and colonic temperature (Tc) was greater in cold-acclimated rats housed at 4 degrees C for 4 weeks (CA) than warm-acclimated controls housed at 24 degrees C for 4 weeks (WA). On the other hand, shivering activity measured at 4 degrees C was less in CA than in WA, while propranolol administration eliminated the difference between these two groups by enhancing shivering in CA. Wet weight and protein content of interscapular brown adipose tissue (IBAT) were greater in CA than in WA. Following cold acclimation, CA were deacclimated at 24 degrees C for 5 weeks. During deacclimation, half of this latter group were forced to run (15 m.min-1 for 1 h) every day (CD-T) while the remaining rats remained sedentary (CD-S). Shivering activity assessed at 4 degrees C 4 weeks after commencing cold deacclimation was significantly less in CD-T than in CD-S and the difference disappeared following propranolol injection. VO2 and Tc responses to NE injection measured 1, 2 and 5 weeks after commencing cold deacclimation did not differ between CD-S and CD-T. Although IBAT weight was lighter in CD-T than in CD-S, its total protein content was not different between the latter two groups of rats. These results suggest that a greater degree of NE-independent nonshivering thermogenesis (NST) is retained in rats that are exercised during the process of deacclimation as compared with animals that are sedentary. This difference in NST would not seem to be directly related to BAT thermogenic capacity.     

Metabolic response to starvation in late pregnant rats. II. Fetal response

Effect of prolonged maternal fasting on the fetal liver and heart glycogen and triglyceride content and on concentration of glucose, urea, uric acid and alpha amino-nitrogen in the amniotic fluid has been studied in rats. The animals were divided into four groups: fed (control), fasted for one day (from 20 to 21 day of pregnancy), fasted for two days (from 19 to 21 day) and fasted for three days (from 18 to 21 day). Maternal fasting for two and three days resulted in reduction in fetal growth. The fetal liver glycogen content was reduced already after one day of fasting, stabilized after two days and then further decreased after three days. The fetal heart glycogen content was reduced only after three days of fasting. The fetal liver triglyceride content increased gradually during the first two days of fasting and then stabilized. The content of triglycerides in the heart was elevated after two and three days of food deprivation. The amniotic fluid glucose concentration decreased after one day of fasting and then stabilized. Fasting did not effect the concentration of the nitrogenous compounds in the amniotic fluid. It is concluded that maternal fasting affects markedly metabolism of energy substrates stored in the fetal liver and the heart and the composition of the amniotic fluid.     

Effect of cold-acclimation on oxygen uptake and glucose production of perfused duckling liver

The control of hepatic metabolism by substrates and hormones was assessed in perfused liver from young Muscovy ducklings. Studies were performed in fed or 24-h fasted 5-week-old thermoneutral (25 degrees C; TN) or cold-acclimated ducklings (4 degrees C; CA) and results were compared with those obtained in rats. Basal oxygen uptake of perfused liver (LVO2) was higher after cold acclimation both in fed (+65%) and 24-h fasted (+29%) ducklings and in 24-h fasted rats (+34%). Lactate (2 mM), the main gluconeogenic substrate in birds, similarly increased LVO2 in both TN and CA ducklings and the effect was larger after fasting. Both glucagon and norepinephrine dose-dependently increased LVO2 in ducklings and rats, but cold acclimation did not improve liver response and liver sensitivity to norepinephrine in ducklings was even reduced in CA animals. Liver contribution to glucagon-induced thermogenesis in vivo was estimated to be 22% in TN and 12% in CA ducklings. Glucagon stimulated gluconeogenesis from lactate in duckling liver and the stimulation was 2.2-fold higher in CA than in TN fasted birds. These results indicate a stimulated hepatic oxidative metabolism in CA ducklings but hepatic glucagon-induced thermogenesis (as measured by LVO2) was not improved. A role of the liver is suggested in duckling metabolic acclimation to cold through an enhanced hepatic gluconeogenesis under glucagon control.     

Clozapine blocks sympathetic and thermogenic reactions induced by orexin A in rat

This experiment tested the effect of clozapine on the sympathetic and thermogenic effects induced by orexin A. The firing rates of the sympathetic nerves to interscapular brown adipose tissue (IBAT), along with IBAT and colonic temperatures were monitored in urethane-anesthetized male Sprague-Dawley rats before and for 5 h after an injection of orexin A (1.5 nmol) into the lateral cerebral ventricle. The same procedure was carried out in rats treated with orexin A plus an intraperitoneal administration of clozapine (8 mg/kg bw), an atypical antipsychotic that is largely used in the therapy of schizophrenia. The same variables were monitored in rats with clozapine alone. A group of rats with saline injection served as control. The results show that orexin A increases the sympathetic firing rate, IBAT and colonic temperatures. Clozapine blocks completely the reactions due to orexin A. These findings suggest that clozapine influences strongly the thermogenic role of orexin A. Furthermore, the remarkable hyperthermic role played by orexin A is confirmed.     

Influence of fasting and hormone deficiency on myocardial glycogen levels in rats.

This study was undertaken because of uncertainties regarding the influence of hormones on myocardial glycogen metabolism of fed and fasted rats. The results indicate that adrenal hormones exert a stabilizing effect on myocardial glycogen levels in fed animals but are not necessary for synthesis to occur. Hypophysectomy eliminates the glycogen increase that occurs from fasting in normal animals while insulin deficiency leads to elevated glycogen stores in both fed and fasted conditions. These findings suggest that changes in myocardial glycogen metabolism are the results of a synergetic relationship between a variety of hormonal and nutritional factors.     

Analysis of metopirone-induced hypertrophy of adrenals in fetal rats encephalectomized in utero

Pregnant rats were treated with 30 mg metopirone (M) each day for 2 days and autopsied on the third day in various gestational periods (Days 18-20, 19-21, and 20-22). Control rats were treated with saline alone (S). The adrenals of intact fetuses in M-treated dams were significantly heavier than those of intact fetuses in S-treated dams in every experimental period. In both M- and S-treated dams, the adrenals of encephalectomized (E) fetuses were lighter than those of intact littermates. However, in the experimental period of Days 18-20 and 19-21, the adrenals of E fetuses in M-treated dams were slightly but significantly heavier than those of similar E fetuses in S-treated dams. In contrast, in the experimental period Days 20-22, there was no significant difference in the weight of adrenals of E fetuses of M- and S-treated dams. These changes in fetal adrenal weight were reflected histologically in parallel changes in the size of adrenocortical cells. The observations suggest that the fetal adrenal hypertrophy following maternal treatment with metopirone can occur to some extent independent of the fetal brain, but that close to the end of gestation the hypertrophy occurs completely under the control of the fetal brain.     

DOPA, dopamine, and DOPAC concentrations in the rat gastrointestinal tract decrease during fasting

The aim of the present study was to test the hypothesis that 3, 4-dihydroxyphenylalanine (DOPA) and dopamine (DA) in the gastrointestinal tract are to a large extent of exogenous origin and derived from food. Tissue concentrations of norepinephrine (NE), epinephrine (Epi), DA, DOPA, and 3,4-dihydroxyphenylacetic acid (DOPAC), as measured by reverse-phase HPLC with electrochemical detection, were studied in fed and 4-day-fasted Wistar rats as well as in sympathectomized and adrenodemedullated rats. Sympathectomy and adrenal demedullectomy decreased tissue concentrations of NE and Epi, respectively, but had no effect on the level of tissue DOPA. Large amounts of DOPA and DA were present in the gastrointestinal tract. Fasting decreased DOPA and DA in the stomach and DOPA concentrations in the quadriceps muscle but no concentrations in other organs. DOPAC in the heart decreased both in response to sympathectomy and to fasting, whereas DOPAC decreased in plasma after fasting and in skeletal muscle after sympathectomy. We conclude that the food content of DOPA and DA is of major importance for the metabolism of DA and, thus, for the dopamine-sulfate content in the gastrointestinal tract and in plasma. The decrease in muscle DOPA after fasting may be explained by less insulin being available during fasting for stimulation of DOPA uptake in the muscle depot. DOPAC in the organism seems to be of a dual origin, derived partly from DA in the food and partly from DA synthesized in sympathetic nerves.     

Plasma free and sulphated catecholamines after ultra-long exercise and recovery

We investigated the early and late effects of two types of ultra-long exercise on sympatho-adrenal and dopaminergic activity. With this aim both free and sulphoconjugated plasma catecholamines (CA), noradrenaline (NA), adrenaline (A), and dopamine (DA) were determined in two groups of athletes immediately after completion of 24-h running or a 10-h triathlon and on recovery during the next 1-3 days. Both races stimulated the sympathetic activity, but differences were observed in the CA pattern: the 24-h run induced a marked elevation of free and sulphoconjugated NA (+175% and +180%, respectively) but failed to alter significantly A and DA levels. The triathlon challenge increased the three conjugated CA (NA sulphate +350%; A sulphate +110%; DA sulphate +270%) and to a lesser extent free CA (NA +45%; A +30%). On the first post-exercise morning, a sustained intense noradrenergic activity was still present in the 24 h-runners, as evidenced by the large increase in free and sulphated NA levels (+140% and +100%, respectively). Such a prolonged activity was also indicated after completion of the triathlon, by the increase of NA sulphate (+140%) observed on the 1st recovery day. However, after the triathlon there was a decreased release of A from the adrenal medulla for several days. These data show that both types of ultralong exercise are able to induce for several hours a sustained sympathetic activation during the test and in the recovery period. Furthermore, the study shows that plasma conjugated CA may provide delayed and cumulative indexes of sympathetic activation, complementary to the instantaneous markers such as free CA.     

Evidence for degradation of intracellular protein in liver lysosomes of fasted rats

Evidence that intracellular protein degradation occurs in lysosomes has been indirect and derived from liver perfusion (1) or the inhibitor studies (2,3). We report here that liver lysosomes of greater purity are obtained from fed rats than from fasted rats. Lysosomes of less purity may contain an enlarged pool of partially degraded intracellular protein; on the other hand, less purity could be due to less marker enzyme, NAβGase. Measurements of NAβGase activity and lysosomal protein of rat livers showed that both NAβGase and lysosomal protein increased upon fasting but protein more so (3.5 and 6.5x, in 2 days). The increase in lysosomal protein is direct evidence that liver lysosomes are involved in intracellular protein degradation during fasting of rats.     

The effect of corticotrophin on liver-type lipase activity in adrenals, liver and high-density lipoprotein subfractions in the rat

Hypercortisolism was induced in rats by the administration of a corticotrophin analogue (Synacthen depot). The effect of this treatment during different periods was studied in normally fed and overnight-fasted rats. The activity of liver-type lipases, i.e., of lipases similar to the heparin-releasable lipase of rat liver (liver lipase), was determined in the adrenal gland and in the liver. Short-term (16 h) treatment had no effect on the lipase activity in the adrenal gland. During prolonged treatment, however, the lipase activity rose to 600-700% of control values in 10 days and from then on remained constant. The effect was similar in fed and overnight-fasted rats. The lipase activity in the liver decreased upon Synacthen administration. In the fed rats a decrease of 25% of the initial value was found after 16 h, 40% after 3 days and 50% after 20 days of treatment. In overnight-fasted rats the lowering of the lipase activity was less marked than in fasted controls. Serum lipid levels and high-density lipoprotein (HDL) subclass concentrations were also measured. The cholesterol concentration in the lipoproteins with a density greater than 1.050 g/ml (HDL) was elevated in rats treated for 3-20 days. If the rats were treated for longer than 10 days, overnight fasting led to a normalization of the HDL-cholesterol levels. After separation of the HDL into two subfractions, a relatively 'light' apolipoprotein E-rich fraction and a more 'heavy' apolipoprotein A-I-rich fraction, in fed and fasted animals treated with Synacthen for 3 days both HDL subfractions were elevated. After 10 days treatment only the apolipoprotein A-I-rich HDL fraction was still enhanced in both fed and fasted rats.     

Increased thermoregulatory responsiveness in cold adapted but not in hyperthyroid hypermetabolic rats

Cold-adapted (CA) rats, unlike non-adapted (NA) ones, give exaggerated metabolic response to acute cold exposure, with paradoxical "overshoot" core temperature (Tc) rise in the cold, and they also give enhanced hyperthermia to central injection of prostaglandin E1 (PGE1). The adaptation-dependent differences might be explained either by the high thermogenic capacity of peripheral tissues in CA rats or by differences in the central processing of regulatory signals. If high tissue metabolism sufficiently explains the extreme responses of CA animals, other hypermetabolic states (with high resting metabolic rate, RMR), e.g. hyperthyroidism, should also be accompanied by enhanced reactions. In the present study thermoregulatory responses to acute cold exposure or to PGE1 were compared in hypermetabolic CA, similarly hypermetabolic thyroxine-treated (T4) and control non-hypermetabolic NA rats (mean RMR = 8.12, 8.47 and 6.03 W kg(-1), respectively). Cold exposure was followed by paradoxical core temperature (Tc) rise of 0.5 to 0.7 degrees C only in CA rats, but by Tc fall (0.8 to 2.1 degrees C) in NA and T4 animals. Identical central stimuli (PGE1) induced larger elevations of Tc and metabolic rate in CA rats than in similarly hypermetabolic T4 or in non-hypermetabolic NA animals (mean Tc rise of 1.9 degrees C in CA vs. 0.9 degrees C in T4 and 1.0 degrees C in NA rats). Vasodilatation thresholds were also similar in NA and T4, but lowered in CA animals. A hypermetabolic status, per se, does not seem to explain the enhanced thermoregulatory responsiveness of CA animals, adaptation-induced central regulatory changes may be more important for the "overshoot" phenomenon.