Reingestion of feces in rodents and its daily rhythmicity

Summary The ingestion of feces is widespread among rodent species and is an extensively employed component of the repertoire of feeding behaviors in some species. Coprophagy is thus a significant consideration in the nutrition and dietary ecology of many rodents. As certain fecal pellets pass from the anus, they are taken up directly into the mouth, chewed, and swallowed. The nocturnally active herbivorous kangaroo rat Dipodomys microps ingests about 1/4 of the feces it produces daily and the daily pattern of reingestion shows a consistent rhythm. For about 8 h of the daytime, during the non-foraging, resting phase of the day, D. microps reingests all fecal pellets produced; during the remainder of the day it leaves all feces produced. The reingested feces contain more nitrogen and water, and less inorganic ions than the non-reingested feces. The extent of reingestion varies among rodent species in relation to diet, and coprophagy is more important in the more herbivorous species. The granivorous kangaroo rat D. merriami ingests feces rarely. The herbivorous vole Microtus californicus ingests about 1/4 of its feces, as does D. microps. However, in contrast to D. microps, M. californicus shows a series of rhythmic, short-term (one to several hour duration) alternations between reingestion and non-reingestion during the course of the day and night. This pattern correlates with the pattern of foraging in M. californicus, which extends over both night and day.     

Immunocytochemical evidence for the presence of an albumin-like substance in the rat pineal gland

Summary Two rabbits and two guinea pigs were immunized with arginine vasotocin (AVT) conjugated to bovine albumin with glutaraldehyde. Only one preparation of antiserum (anti-G 7) was of value. Anti-G 7 immunochemically defined the same rat pineal cells previously reported as presumptive AVT cells. However, absorption of anti-G 7 with bovine albumin inhibited the staining of the pineal cells demonstrating that they contained an albumin-like substance. Positive immunochemical staining of the rat pars nervosa suggested that anti-G 7 contained antibodies able to react with arginine vasopressin (AVP). Loss of a positive reaction in the posterior lobe on absorption of anti-G 7 with AVT or AVP confirmed this. However, the addition of AVT to anti-G 7 failed to inhibit the immunochemical staining of the pineal cells. This study reports the presence of an albumin-like substance in pineal cells previously described as presumptive AVT cells, and discusses possible explanations for the inability of anti-G 7 to recognize immunocytochemically the native AVT molecule. Confirmation of AVT in the pineal gland by immunocytochemistry must await the availability of more specific antisera.     

Mass spectrometric identification and quantification of 5-methoxytryptophol in quail retina

The occurrence of 5-methoxytryptophol (5-MTL) in the quail retina was investigated by capillary column gas chromatography/mass spectrometry/selected ion monitoring using a deuterated internal standard. Based on ion intensity ratios in the mass spectra of pentafluoropropionyl and heptafluorobutyryl derivatives of 5-MTL and deuterated 5-MTL, 5-MTL was unequivocally identified in the quail retina. Similar to the circadian rhythm of retinal melatonin, retinal 5-MTL also exhibited a diurnal variation with high levels at mid-dark. However, no significant correlation between the diurnal levels of 5-MTL and melatonin was observed in the quail retina at mid-light or mid-dark.     

Type-II thyroxine 5'-deiodinase is present in the rat pineal gland

Thyroxine-5'-deiodinase has been identified in the rat pineal gland. The characteristics of the enzyme are compatible with a Type-II deiodinase which is tissue-specific and presumably related to generating a local action of thyroid hormone. Our data suggest there may be a previously unrecognized role of thyroid hormone in the regulation of pineal activity.     

Circadian rhythmicity in the rat exocrine pancreas: chronomorphological patterns

Circadian rhythmicity of the structural morphometric model of rat endocrine pancreas has been studied in 24 Wistar female rats, four months old, kept in LD 12:12. The following parameters were evaluated: the volume fractions of nucleus and cytoplasm of exocrine cells, the size distribution and number in unit tissue volume of acinar cell nuclei, the mean nuclear diameter, the shape coefficient of glandular acini (that is the ratio acinar area/perimeter2 which indicates the shifting of structures from circularity). A statistically significant circadian rhythm was demonstrated for the shape coefficient of glandular acini. Results obtained in the present experiment are compared with data recorded in a previous study.     

Methionine adenosyltransferase:adrenergic-cAMP mechanism regulates a daily rhythm in pineal expression

(S)-adenosylmethionine (SAM) is a critical element of melatonin synthesis as the methyl donor in the last step of the pathway, the O-methylation of N-acetyl 5-hydroxytryptamine by hydroxyindole-O-methyltransferase. The activity of the enzyme that synthesizes SAM, methionine adenosyltransferase (MAT), increases 2.5-fold at night in the pineal gland. In this study, we found that pineal MAT2A mRNA and the protein it encodes, MAT II, also increase at night, suggesting that the increase in MAT activity is caused by an increase in MAT II gene products. The night levels of MAT2A mRNA in the pineal gland were severalfold higher than in other neural and non-neural tissues examined, consistent with the requirement for SAM in melatonin synthesis. Related studies indicate that the nocturnal increase in MAT2A mRNA is caused by activation of a well described neural pathway that mediates photoneural-circadian regulation of the pineal gland. MAT2A mRNA and MAT II protein were increased in organ culture by treatment with norepinephrine (NE), the sympathetic neurotransmitter that stimulates the pineal gland at night. NE is known to markedly elevate pineal cAMP, and here it was found that cAMP agonists elevate MAT2A mRNA levels by increasing MAT2A mRNA synthesis and that drugs that block cAMP activation of cAMP dependent protein kinase block effects of NE. Therefore, the NE-cAMP dependent increase in pineal MAT activity seems to reflect an increase in MAT II protein, which occurs in response to cAMP-->protein kinase-dependent increased MAT2A expression. The existence of this MAT regulatory system underscores the importance that MAT plays in melatonin biogenesis. These studies also point to the possibility that SAM production in other tissues might be regulated through cAMP.     

Search for seasonal rhythmicity of pineal melatonin production in rats under constant laboratory conditions: spectral chronobiological analysis, and relation to solar and geomagnetic variables

Earlier we reported that in a number of experiments pineal melatonin production in rats under constant laboratory conditions displayed seasonal rhythms but subsequently were not always able to confirm this. Since there was no indication under which conditions such rhythms may be present, we performed four consecutive identical experiments with untreated female Sprague-Dawley rats within the same animal room during 1997-2006. Nocturnal urine samples (19-23, 23-3, 3-7?h) were collected at monthly intervals over 494-658?d with 12 animals each in experiments I and II (1997-1999, 1999-2000), 30 animals in experiment III (2002-2004), and 15 in experiment IV (2005-2006). 6-Sulfatoxymelatonin (aMT6s) was measured by ELISA. The excreted aMT6s at each time interval as well as total nocturnal aMT6s-excretion (19-7?h) was submitted to standard statistical analyses as well as to a spectral chronobiological analysis to determine the period lengths of the components involved which was followed by processing with the single cosinor method. Seasonal rhythm components (circannual period length: 360 ± 60?d) were detected in experiment III (2002-2004) for the overall nocturnal excretion as well as for two sub-intervals (23-3 and 3-7?h) and in one night interval of experiment II (23-3?h). Multiple components with mostly short period lengths of around 100?d and some long ones of 500-650?d were found in the other experiments. Systematic MESOR and amplitude variations were observed during the experiments, being highest in experiment II (19-7?h, also 23-3?h and 3-7?h) and lowest in experiments I and IV. These results illustrate that seasonal melatonin rhythms are not a general phenomenon in female laboratory rats indicating an involvement of unknown environmental cues. As an extension of our earlier hypothesis regarding a seasonal Zeitgeber function of the horizontal intensity H of the geomagnetic field showing circannual variations, we assume further modulation by the 11-yrs' sunspot cycle which leads to geomagnetic disturbances and could facilitate seasonal aMT6s rhythmicity during specific years. (Author correspondence: christian.bartsch@uni-tuebingen.de ).     

Age-related differences in serum melatonin and pineal NAT activity and in the response of rat pineal to a 50-Hz magnetic field.

In a previous study we have shown that exposure to a 50-Hz sinusoidal magnetic field decreased serum melatonin concentration and pineal enzyme activities in young rats (9 weeks). In the present study we looked for the effect of a magnetic field of 100 microT on serum melatonin and pineal NAT activity in aged rats and compared them to young rats. We hypothesized that aging may change sensitivity of rats to a magnetic field. Two groups of Wistar male rats [aged rats (23 months) and young rats (9 weeks)] were exposed to 50-Hz magnetic fields of 100 microT for one week (18h/day). The animals were kept under a standard 12:12 light: dark cycle with a temperature of 25 degrees C and a relative humidity of 45 to 50%. Control (sham-exposed) animals were kept in a similar environment but without exposure to a magnetic field. The animals were sacrificed under red dim light. Serum melatonin concentration and pineal N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT) activities were studied. Our results showed that sinusoidal magnetic fields altered the production of melatonin (28% decrease; P <0.05) through an inhibition of pineal NAT activity (52% decrease; P <0.05) in the young rats whereas no effect was observed in aged ones. On the other hand, when comparing data from control animals between young and aged rats, we observed that serum melatonin level and NAT activity, but not HIOMT activity, decreased in aged rats (decrease by about 38% and 36% respectively). Our data strongly suggest that old rats are insensitive to the magnetic field.     

Negative correlation of age and the levels of pineal melatonin, pineal N-acetylserotonin, and serum melatonin in male rats

Pineal concentrations of N-acetylserotonin and melatonin and serum levels of melatonin were studied in 3-wk-old (prepubertal), 8-wk-old (adult), and 17-mo-old (senile) male rats. They were adapted to a photoperiod of 12 h light/12 h darkness for a minimum of 1 wk and killed at mid-light and mid-dark. Melatonin and N-acetylserotonin were determined by radioimmunoassay. The concentrations of pineal N-acetylserotonin and melatonin were high in the dark period and low in the light period. Statistical analysis indicated that pineal N-acetylserotonin and melatonin levels per 100 gm body weight declined with age. Similarly, serum melatonin demonstrated diurnal changes in all the age groups studied. In addition, there was a significant reduction in the levels of serum melatonin with age. The parallel patterns of decrease in pineal and serum melatonin levels with age suggest a decline in pineal secretion of melatonin in the older animals.     

Effects of carbohydrate and protein administration on rat tryptophan and 5-hydroxytryptamine: differential effects on the brain, intestine, pineal, and pancreas

We compared the acute effects of intragastric administration of protein and carbohydrate on tryptophan and 5-hydroxytryptamine (5HT) in rat brain, pineal, intestine, and pancreas. Protein decreased and carbohydrate increased brain indoles relative to water-infused controls. These effects were due to competition between the large neutral amino acids for entry into the brain. This competition does not exist in the pineal. The macronutrients had no effect on pineal tryptophan metabolism. In the intestine, protein resulted in higher tryptophan levels as compared to controls, owing to absorption of tryptophan in the protein. However intestinal 5HT levels were influenced by factors other than precursor availability. Pancreatic indoles were affected in a similar manner to the brain indoles. Competition between the large neutral amino acids for entry into the pancreas was also indicated by the finding that valine administration lowered brain and pancreatic tryptophan, but not the levels in the intestine and pineal. It remains to be seen whether the decrease in pancreatic 5HT after a protein meal and the increase after carbohydrate modulate the release of insulin and glucagon.     

Entrainment of daily serum gonadotropin cycles in the goldfish to photoperiod, feeding, and daily thermocycles

Female fish were kept under 16L:8D/20 degrees C in November and April and the onset of light and/or feeding times were shifted by several hours in the experimental groups. Photoperiod and feeding entrained significant fluctuations in serum gonadotropin hormone (GTH) levels when the onset of light and the first daily feeding were 4 hr apart, but not when they were 10 hr apart. Fish were subjected to 16L:8D for 14-16 days in February, and to either a constant warm (20 degrees C) or a diurnal sinusoidal (12-20 degrees C) temperature regime, the warmth being imposed during photophase or scotophase. While relatively high, uniform serum GTH levels were found throughout the 24-hr period in fish subjected to constant warmth, warm temperature during the day promoted fluctuations in serum GTH levels, and warmth during night resulted in relatively low, uniform serum GTH levels.     

Synaptic ribbons in the pineal organ of the goldfish: Circadian rhythmicity and the effects of constant light and constant darkness

Summary Synaptic ribbons in photoreceptor cells of the goldfish pineal organ undergo significant daily changes in their length, distance from the plasma membrane, and number per unit area of pineal end-vesicle. The rhythms persist in fish exposed to constant darkness. Constant light abolishes the rhythms in length and distance of synaptic ribbons from the plasmalemma, but has little effect on numerical changes over a 24-h cycle. These findings suggest that synaptic ribbons in the pineal organ of lower vertebrates might be useful as indicators of metabolic activity.     

Early development of circadian rhythmicity in the suprachiamatic nuclei and pineal gland of teleost,flounder (Paralichthys olivaeus), embryos

Circadian rhythms enable organisms to coordinate multiple physiological processes and behaviors with the earth's rotation. In mammals, the suprachiasmatic nuclei (SCN), the sole master circadian pacemaker, has entrainment mechanisms that set the circadian rhythm to a 24‐h cycle with photic signals from retina. In contrast, the zebrafish SCN is not a circadian pacemaker, instead the pineal gland (PG) houses the major circadian oscillator. The SCN of flounder larvae, unlike that of zebrafish, however, expresses per2 with a rhythmicity of daytime/ON and nighttime/OFF. Here, we examined whether the rhythm of per2 expression in the flounder SCN represents the molecular clock. We also examined early development of the circadian rhythmicity in the SCN and PG. Our three major findings were as follows. First, rhythmic per2 expression in the SCN was maintained under 24 h dark (DD) conditions, indicating that a molecular clock exists in the flounder SCN. Second, onset of circadian rhythmicity in the SCN preceded that in the PG. Third, both 24 h light (LL) and DD conditions deeply affected the development of circadian rhythmicity in the SCN and PG. This is the first report dealing with the early development of circadian rhythmicity in the SCN in fish.