Geneviève Bourdel

Circadian feeding pattern in pregnant rats fed three levels of protein.

Food intake was measured at regular intervals over 24 h in pregnant and non-pregnant female rats fed diets of different protein content: 10, 16 and 32%. During the course of pregnancy, a first period of hyperphagia was observed (days 2-12) irrespective of the composition of the diet. A second phase of hyperphagia occurred later (days 16-19) which was more marked with the better balanced diet (16% protein). During the first half of pregnancy, the increase in intake occurred principally at the beginning of the night (compensatory reaction). Later on, the stimulation extended to the last part of the night (anticipatory reaction). The nocturnal predominance of feeding activity was maintained in pregnant females in spite of their increased metabolic requirements.

Regulation of hepatic synthesis of proteins by the chronology of protein ingestion

Circadian variations in liver protein synthesis were were assessed in control rats fed a mixed 10% protein diet and in rats fed proteins as a separate meal either at 09:00 (SF 09) or at 21:00 (SF 21) and provided with a protein-free diet ad libitum. Protein synthesis was measured by incorporation of labelled leucine over a short period of time (15 min) at time-points regularly spaced over 24 h. In controls, the circadian variations observed were of moderate amplitude (from 2.75 mg/h per g at 09:00 to 5.77 mg/h per g at 06:00) correlated with increased protein and RNA contents of the liver. In separately fed animals ingestion of the protein meal triggered a 300% increase in protein synthesis within 1 h while the feeding pattern was unaltered. In the SF 09 group, high synthetic activity was not followed by an increase of hepatic protein content while hepatic urea concentrations were sharply increased and glucogenic amino acid pools were greatly depleted. It is suggested that the high influx of amino acids consecutive to the absorption of the dietary proteins is the key factor stimulating protein synthesis, while synchronisation with the energetic metabolism controls the degree of degradation. The possible involvement of variations in the insulin to glucagon ratio is discussed.

Circadian variations of A-mediated transport in rat-liver plasma membrane vesicles

Among the 3 transport systems for neutral amino acids known to operate at the hepatic level, only system A has been shown to be responsive to hormonal and nutritional stimuli. The inductive effect of glucagon [ 151, insulin [2,5,8], corticoids [ 1,9], and catecholamines [ 1,101 is now well documented. Direct short-term induction of stimulated A-mediated transport by amino acid deprivation and reversal by amino acid supplementation, has been shown in cultures hepatocytes [ 111. Hormonal and/or nutritional treatments applied in vivo, induce changes in hepatic A-mediated transport that can be readily demonstrated in vitro at the cellular level [ 1,12,13]. Rats kept on regular light-dark schedules exhibit distinct rhythms of hormonal secretions and ingest most of their food (70%) during the night. This behavior results in alternate phases of low (light-phase) and high (dark-phase) portal aminoacidemia [14]. Circadian changes in hepatic Amediated transport can thus be expected 1151. Here, amino acid transport activity was measured in plasma-membrane vesicles from the liver of rats fed ad libitum a low casein diet and killed at different time-points of the light-dark cycle. This technique has the advantage of using natural amino acids as substrate. Proline was chosen as substrate because, more than alanine [ 16,171, it is preferentially transported through system A in hepatocytes. 53% starch, 20% sucrose, 8% peanut oil, 2% cellulose, 4% mineral mixture, and 1% vitamin mixture. After 3 weeks, they were killed by decapitation in groups of 6 at regularly spaced time-points: 4 groups during the light-phase (3,6,9 and 12 h exposure to light), 4 groups during the dark-phase (3,6,9 and 12 h exposure to darkness).

Induction of multiple forms of tyrosine aminotransferase by amino acid mixtures of different compositions

Hepatic tyrosine aminotransferase (EC 2.6.1.5) was induced in rats by intubation of amino acid mixtures (complete or tryptophan-free). Enzyme activity was increased 4-fold by the complete mixture and 8-fold by the tryptophan-free mixture. The enzyme was analyzed by chromatography on CM-Sephadex. Chromatographic patterns were characteristic of the type of inducer rather than of the chronology of the induction cycle: after induction by the complete amino acid mixture the three forms of the enzyme were equally increased whereas after induction by the tryptophan-free mixture Form I was preferentially increased.

Na(+)-dependent transport of alanine and serine by liver plasma-membrane vesicles from rats fed a low-protein or a high-protein diet.

Plasma-membrane vesicles prepared from the liver of rats fed either a low-(LP) or a high-protein (HP) diet exhibited Na(+)-dependent active transport of alanine and serine. The process gave apparent kinetic parameters compatible with a single saturable component for both amino acids. Na,K-ATPase (EC 3.6.1.37), marker of the basolateral domain of the hepatocyte plasma-membrane, was chosen as reference for the expression of amino acid transport in vesicle preparations. The high-protein diet induced a significant increase in liver Na,K-ATPase activity also found in corresponding plasma-membrane preparations, in parallel with an increase in the capacity towards amino acid transport. This suggests that in rats fed the high protein diet, transcellular Na+ exchange, although increased, remains well balanced. N-Methylaminoisobutyric acid (MeAIB), due to its poor velocity, proved unsuitable to distinguish between systems A and ASC in the experimental model. Comparing Na(+)- and Li(+)-driven transport, a family of carriers with strict Na(+)-dependency (A-like) was evidenced in LP vesicles but not in HP vesicles. The sensitivity to the lowering of the pH from 7.5 to 6.5 in the external medium was similar in both type of vesicles when Na+ was the driving ion. In the HP vesicles the Li(+)-tolerant, pH-insensitive component (ASC-like) was increased in parallel with overall Na(+)-dependent transport. These functional properties suggest that the carriers involved in the stimulation of transport in HP vesicles are composite in nature. Increasing concentrations of an amino acid mixture mimicking the changes of portal aminoacidemia inhibited the transport of alanine and of serine. The degree of inhibition was correlated with the relative concentration of substrate and was independent of the nutritional treatment.