Early weaning accelerates the differentiation of mucous neck cells in rat gastric mucosa: Possible role of TGF alpha/EGFR

The development of the gastric mucosa is controlled by hormones, growth factors and feeding behavior. Early weaning (EW), which means the abrupt interruption of suckling, increases proliferation and differentiation in the rat gastric epithelium. Transforming growth factor alpha(TGF alpha) is secreted in the stomach, binds to the epidermal growth factor receptor( EGFR) and may control cell proliferation, differentiation and migration. Here, we investigated the influence of suckling-weaning transition on the differentiation of mucous neck cells in the stomach and its association to the expression of TGF alpha and EGFR. Fifteen-day-old Wistar rats were divided into two groups: suckling( control), in which pups were kept with the dam, and early weaning( EW), in which rats were separated from their mother and fed with hydrated powdered chow. TGF alpha and EGFR levels were increased at 18 days in EW animals compared to control ones (p<0.05). Histochemical reactions with Periodic Acid-Schiff reagent+Alcian Blue or Bandeiraea simplicifolia II lectin were used to stain the mucous neck cells and showed an increase in this cell population throughout EW, which was more pronounced at 17 days when compared to suckling pups (p<0.05). These morphological results were confirmed by RT-PCR for mucin 6. The levels of mucin 6 mRNA were higher in EW animals from the 16th to the 18th day(1-3 days post-weaning) when compared to the respective control group. Inhibition of EGFR through AG1478 administration to EW animals prevented the expansion of mucous neck cell population induced by EW (p<0.05). Therefore, early weaning up regulated TGF alpha/EGFR expression and induced differentiation of mucous neck cells. Moreover, we showed that EGFR takes part in the maturation of this cell population. We conclude that regular suckling-weaning transition is crucial to guarantee the development of the gastric mucosa. (C) 2009 International Society of Differentiation. Published by Elsevier Ltd. All rights reserved.

Eag 1, Eag 2 and Kcnn3 gene brain expression of isolated reared rats

The Eag1 and Eag2, voltage-dependent potassium channels, and the small-conductance calcium-activated potassium channel (Kcnn3) are highly expressed in limbic regions of the brain, where their function is still unknown. Eag1 co-localizes with tyrosine hydroxilase enzyme in the substantia nigra and ventral tegmental area. Kcnn3 deficiency leads to enhanced serotonergic and dopaminergic neurotransmission accompanied by distinct alterations in emotional behaviors. As exposure to stress is able to change the expression and function of several ion channels, suggesting that they might be involved in the consequences of stress, we aimed at investigating Eag 1, Eag2 and Kcnn3 mRNA expression in the brains of rats submitted to isolation rearing. As the long-lasting alterations in emotional and behavioral regulation after stress have been related to changes in serotonergic neurotransmission, expressions of serotonin Htr1a and Htr2a receptors in male Wistar rats` brain were also investigated. Rats were reared in isolation or in groups of five for nine weeks after weaning. Isolated and socially reared rats were tested for exploratory activity in the open field test for 5 min and brains were processed for reverse-transcription coupled to quantitative polymerase chain reaction (qRT-PCR). Isolated reared rats showed decreased exploratory activity in the open field. Compared to socially reared rats, isolated rats showed reduced Htr2a mRNA expression in the striatum and brainstem and reduced Eag2 mRNA expression in all examined regions except cerebellum. To our knowledge, this is the first work to show that isolation rearing can change Eag2 gene expression in the brain. The involvement of this channel in stress-related behaviors is discussed.

Effect of inbreeding on growth and reproductive traits of Nellore cattle in Brazil

The objective of this study was to identify and quantify the influence of F (inbreeding coefficient) on weaning weight (WW), weight gain from weaning to 18 months of age (WG345), finishing visual score (precocity) at 18 months of age, muscling visual score at 18 months of age (MUS), hip height (HH), scrotal circumference at 18 months of age (SC), heifer probability of pregnancy at 14 months of age (PP14), and stayability (STAY) in Brazilian Nellore cattle. The complete pedigree included 417,552 animals born between 1984 and 2007 on 12 farms located in the states of Mato Grosso do Sul, Sao Paulo and Bahia. Following the observation of a statistically significant effect (P<0.05) of the covariates individual inbreeding coefficient (F) and maternal inbreeding coefficient, regression analysis of each trait, adjusted for all other effects, was performed as a function of the linear and quadratic effect of F and maternal F (when significant). Inbreeding negatively affected all traits studied (P<0.05), except for muscling. A quadratic effect of individual F on WW, WG345, HH and PP14, and a quadratic effect of maternal F on WG345 and HH were observed. Levels of inbreeding higher than 7-11% affected negatively growth and reproductive performance of Nellore cattle. Therefore, inbreeding should be avoided, except for purposes of genetic breeding whose main objective is the fixation of certain alleles in the population. (C) 2010 Elsevier B.V. All rights reserved.

Hormone release and behavior during suckling and milking in Gir, Gir x Holstein, and Holstein cows

There are several different milking management systems in Latin America, because Gir cattle are reputed to be easily stressed and not well adapted to machine-milking. This paper, therefore, provides an overview of hormone release and behavior during suckling and milking in Gir cows and their crossbred offspring. Several experiments were performed to study oxytocin release during exclusive suckling or exclusive hand- and machine-milking, oxytocin, and prolactin release during a mixed suckling-milking system and oxytocin release after weaning. Cortisol concentrations and behavior were also examined. Concentration of oxytocin, released during suckling, and both types of milking were high, but the maximum concentration measured during suckling was significantly greater than that observed during exclusive milking. In the mixed suckling-milking system, the greatest oxytocin and prolactin releases were measured during suckling. Cortisol concentrations measured before, during, and after milking demonstrated that Gir x Holstein and Holstein cows were not stressed. On the other hand, although Gir had greater concentrations of cortisol, the percentage of residual milk for Gir cows was less than for dairy cows exposed to different stressful situations. In general, Gir cows and their crossbred offspring adapted to machine-milking, although these breeds can react negatively to milkers. Gir, Gir x Holstein, and Holstein cows all had similar cortisol levels during and after milking.

Assessment of DGAT1 and LEP gene polymorphisms in three Nelore (Bos indicus) lines selected for growth and their relationship with growth and carcass traits

The aim of this study was to analyze LEP and DGAT1 gene polymorphisms in 3 Nelore lines selected for growth and to evaluate their effects on growth and carcass traits. Traits analyzed were birth, weaning, and yearling weight, rump height, LM area, backfat thickness, and rump fat thickness obtained by ultrasound. Two SNP in the LEP gene [LEP 1620(A/G) and LEP 305(T/C)] and the K232A mutation in the DGAT1 gene were analyzed. The sample consisted of 357 Nelore heifers from 2 lines selected for yearling weight and a control line, established in 1980, at the Estacao Experimental de Zootecnia de Sertaozinho (Sertaozinho, Brazil). Three genotypes were obtained for each marker. Differences in allele frequencies among the 3 lines were only observed for the DGAT1 K232A polymorphism, with the frequency of the A allele being greater in the control line than in the selected lines. The DGAT1 K232A mutation was associated only with rump height, whereas LEP 1620(A/G) was associated with weaning weight and LEP 305(T/C) with birth weight and backfat thickness. However, more studies, with larger data sets, are necessary before these makers can be used for marker-assisted selection.

Postnatal fluoxetine treatment affects the development of serotonergic neurons in rats

The goal of the present study was to investigate morphological changes in the serotonergic neurons/terminals in the dorsal (DR) and median (MnR) raphe nuclei and on the hippocampal dentate gyrus (DG) in neonatal rats treated from the 1st to the 21st postnatal day with fluoxetine (10 mg/kg sc, daily) or drug vehicle (0 9% saline 1 ml/kg). The results show that postnatal chronic treatment with fluoxetine promoted. (1) a smaller body weight increase during the pre-weaning period; (2) smaller number of 5-HT neurons in the DR, (3) smaller 5-HT neuronal cell bodies (area, perimeter and diameter) in the DR and the MnR and (4) diminished serotonergic terminals in the DG. These data suggest that the development of the serotonergic system was impaired and that early exposure to fluoxetine damaged the morphology of 5-HT neurons in young adult rats While these findings are consistent with other work, more studies are needed to better clarify the effects of postnatal chronic treatment with fluoxetine on the serotonergic system and, consequently, on the functions modulated by serotonin (C) 2010 Elsevier Ireland Ltd All rights reserved

Prolonged consumption of soy or fish-oil-enriched diets differentially affects the pattern of hypothalamic neuronal activation induced by refeeding in rats

We used c-Fos immunoreactivity to estimate neuronal activation in hypothalamic feeding-regulatory areas of 3-month-old rats fed control or oil-enriched diets (soy or fish) since weaning. While no diet effect was observed in c-Fos immunoreactivity of 24-h fasted animals, the acute response to refeeding was modified by both hyperlipidic diets but with different patterns. Upon refeeding, control-diet rats had significantly increased c-Fos immunoreactivity only in the paraventricular hypothalamic nucleus (PVH, 142%). In soy-diet rats, refeeding with the soy diet increased c-Fos immunoreactivity in dorsomedial hypothalamic nucleus (DMH, 271%) and lateral hypothalamic area (LH, 303%). Refeeding fish-diet rats with the fish diet increased c-Fos immunoreactivity in PVH (161%), DMH (177%), VMH (81%), and ARC (127%). Compared to the fish-diet, c-Fos immunoreactivity was increased in LH by the soy-diet while it was decreased in ventromedial hypothalamic nucleus (VMH) and arcuate hypothalamic nucleus (ARC). Based on the known roles of the activated nuclei, it is suggested that, unlike the fish-diet, the soy-diet induced a potentially obesogenic profile, with high LH and low VMH/PVH activation after refeeding.

Can physical exercise during gestation attenuate the effects of a maternal perinatal low-protein diet on oxygen consumption in rats?

A protocol of physical exercise, based on maximal oxygen uptake ((V) over dot(O2max)), for female rats before and during pregnancy was developed to evaluate the impact of a low-protein diet on oxygen consumption during gestation and growth rate of the offspring. Virgin female Wistar rats were divided into four groups as follows: untrained (NT, n = 5); trained (T, n = 5); untrained with low-protein diet (NT+LP, n = 5); and trained with low-protein diet (T+LP, n = 5). Trained rats were submitted to a protocol of moderate physical training on a treadmill over a period of 4 weeks (5 days week(-1) and 60 min day(-1), at 65% of (V) over dot(O2max)). At confirmation of pregnancy, the intensity and duration of the exercise was reduced. Low-protein groups received an 8% casein diet, and their peers received a 17% casein diet. The birthweight and growth rate of the pups up to the 90th day were recorded. Oxygen consumption ((V) over dot(O2)), CO(2) production and respiratory exchange ratio (RER) were determined using an indirect open-circuit calorimeter. Exercise training increased. (V) over dot(O2max) by about 20% when compared with the initial values (45.6 +/- 1.0 ml kg(-1) min(-1)). During gestation, all groups showed a progressive reduction in the resting (V) over dot(O2) values. Dams in the NT+LP group showed lower values of resting (V) over dot(O2) than those in the NT group. The growth rate of pups from low-protein-fed mothers was around 50% lower than that of their respective controls. The T group showed an increase in body weight from the 60th day onwards, while the NT+LP group presented a reduced body weight from weaning onwards. In conclusion, physical training attenuated the impact of the low- protein

Glucose metabolism by lymphocytes, macrophages, and tumor cells from Walker 256 tumor-bearing rats supplemented with fish oil for one generation

Here we investigated the effect of lifelong supplementation of the diet with coconut fat (CO, rich in saturated fatty acids) or fish oil (170, rich in n-3 polyunsaturated fatty acids) on tumor growth and lactate production from glucose in Walker 256 tumor cells, peritoneal macrophages, spleen, and gut-associated lymphocytes. Female Wistar rats were supplemented with CO or FO prior to mating and then throughout pregnancy and gestation and then the male offspring were supplemented from weaning until 90 days of age. Then they were inoculated subcutaneously with Walker 256 tumor cells. Tumor weight at 14 days in control rats (those fed standard chow) and CO supplemented was approximately 30 g. Supplementation of the diet with FO significantly reduced tumor growth by 76%. Lactate production (nmol h(-1) mg(-1) protein) from glucose by Walker 256 cells in the group fed regular chow (W) was 381.8 +/- 14.9. Supplementation with coconut fat (WCO) caused a significant reduction in lactate production by 1.6-fold and with fish oil (WFO) by 3.8-fold. Spleen lymphocytes obtained from W and WCO groups had markedly increased lactate production (553 +/- 70 and 635 +/- 150) when compared to non-tumor-bearing rats (similar to 260 +/- 30). FO supplementation reduced significantly the lactate production (297 +/- 50). Gut-associated lymphocytes obtained from W and WCO groups increased lactate production markedly (280 +/- 31 and 276 +/- 25) when compared to non-tumor-bearing rats (similar to 90 +/- 18). FO supplementation reduced significantly the lactate production (168 +/- 14). Lactate production by peritoneal macrophages was increased by tumor burden but there was no difference between the groups fed the various diets. Lifelong consumption of FO protects against tumor growth and modifies glucose metabolism in Walker tumor cells and lymphocytes but not in macrophages. Copyright (C) 2008 John Wiley & Sons, Ltd.

High sodium intake enhances insulin-stimulated glucose uptake in rat epididymal adipose tissue

Objective: This study investigated the effect of different sodium content diets on rat adipose tissue carbohydrate metabolism and insulin sensitivity. Methods and Procedures: Male Wistar rats were fed on normal- (0.5% Na+; NS), high- (3.12% Na+; HS), or low-sodium (0.06% Na+; LS) diets for 3, 6, and 9 weeks after weaning. Blood pressure (BP) was measured using a computerized tail-cuff system. An intravenous insulin tolerance test (ivITT) was performed in fasted animals. At the end of each period, rats were killed and blood samples were collected for glucose and insulin determinations. The white adipose tissue (WAT) from abdominal and inguinal subcutaneous (SC) and periepididymal (PE) depots were weighed and processed for adipocyte isolation and measurement of in vitro rates of insulin-stimulated 2-deoxy-d-[H-3]-glucose uptake (2DGU) and conversion of -[U-C-14]-glucose into (CO2)-C-14. Results: After 6 weeks, HS diet significantly increased the BP, SC and PE WAT masses, PE adipocyte size, and plasma insulin concentration. The sodium dietary content did not influence the whole-body insulin sensitivity. A higher half-maximal effective insulin concentration (EC50) from the dose - response curve of 2DGU and an increase in the insulin-stimulated glucose oxidation rate were observed in the isolated PE adipocytes from HS rats. Discussion: The chronic salt overload enhanced the adipocyte insulin sensitivity for glucose uptake and the insulin-induced glucose metabolization, contributing to promote adipocyte hypertrophy and increase the mass of several adipose depots, particularly the PE fat pad.

Opposite effects of fasting on TGF-beta 3 and T beta RI distribution in the gastric mucosa of suckling and early weanling rats

Objective: Our aim was to evaluate the effects of a dietary regimen (suckling or early weaning) and feeding status (fed or fasted) on the distribution of transforming growth factor-beta 3 (TGF-beta 3) and TGF receptor-I (T beta RI) in the gastric epithelium of pups Methods: Wistar rats were used At 15 d, half of the pups were separated from dams and fed with hydrated powered chow On day 17, suckling and early weanling rats were subjected to fasting (17 h). Four different conditions were established. suckling fed and fasted and early weanling fed and fasted At 18 d stomachs were collected under anesthesia and were fixed in 4% formaldehyde for immunohistochemistry The number of immunostained epithelial cells per microscopic field was determined for TGF-beta 3 and T beta RI in longitudinal sections from the gastric mucosa Results: We found that during suckling, fasting reduced the number of immunolabeled cells per field of both molecules when compared with the fed group (P < 0.05), whereas in early weaning, food restriction increased TGF-beta 3 and T beta RI distributions (P < 0.05) We also observed that TGF-beta 3 and T beta RI were more concentrated in parietal cells in the upper gland in suckling pups, whereas after early weaning these were displaced to parietal and chief cells at the bottom of the gland Conclusion: Suckling and early weaning directly influence TGF-beta 3 and T beta RI distributions in the gastric epithelium in response to fasting, such that early weaning anticipates the effects observed in adult rats. Furthermore, the differential concentrations of TGF-beta 3 and T beta RI indicate that they might be important for cell proliferation events in growth control (C) 2010 Elsevier Inc. All rights reserved

EGFR is involved in control of gastric cell proliferation through activation of MAPK and Src signalling pathways in early-weaned rats

Objectives: Early weaning (EW) increases proliferation of the gastric epithelium in parallel with higher expression of transforming growth factor alpha and its receptor epidermal growth factor receptor (EGFR). The primary objective of the present study was to examine involvement of EGFR signalling in regulating mucosal cell proliferation during the early weaning period. Materials and methods: Fifteen-day-old rats were split into two groups: suckling (control) and EW, in which pups were separated from the dam. Animals were killed daily until the 18th day, 3 days after onset of treatment. To investigate the role of EGFR in proliferation control, EW pups were injected with AG1478, an EGFR inhibitor; signalling molecules, proliferative indices and cell cycle-related proteins were evaluated. Results: EW increased ERK1/2 and Src phosphorylation at 17 days, but p-Akt levels were unchanged. Moreover, at 17 days, AG1478 administration impaired ERK phosphorylation, whereas p-Src and p-Akt were not altered. AG1478 treatment reduced mitotic and DNA synthesis indices, which were determined on HE-stained and BrdU-labelled sections. Finally, AG1478 injection decreased p21 levels in the gastric mucosa at 17 days, while no changes were detected in p27, cyclin E, CDK2, cyclin D1 and CDK4 concentrations. Conclusions: EGFR is part of the mechanism that regulates cell proliferation in rat gastric mucosa during early weaning. We suggest that such responses might depend on activation of MAPK and/or Src signalling pathways and regulation of p21 levels.

In vivo effects of TGF beta 1 on the the growth of gastric epithelium in suckling rats

As the content of Transforming Growth Factor-beta (TGF beta) wanes in the milk of lactating rat, an increase in TGF beta is observed in the gastric epithelia concomitant with differentiation of the glands upon weaning. Whereas TGF beta has been shown to inhibit the proliferation of gastrointestinal cells in vitro, its functional significance and mechanisms of action have not been studied in vivo. Therefore, we administered TGF beta 1 (1 ng/g body wt.) to 14-day-old rats in which the gastric epithelium was induced to proliferate by fasting, and determined the involvement of signaling through Smads and the impact on epithelial cell proliferation and apoptosis. After the gavage, we observed the progressive increase of active TGF beta 1 while T beta RII-receptor remained constant in the gastric mucosa. By immunohistochemistry, we showed Smad2/3 increase at 60 min (p < 0.05) and Smad2 phosphorylation/activation and translocation to the nucleus most prominently between 0 and 30 min after treatment (p < 0.05). Importantly, TGF beta 1 inhibited cell proliferation (p < 0.05), which was estimated by BrDU pulse-labeling 12 h after gavage. Lower proliferation was reflected by increased p27(kip1) at 2 h (p < 0.05). Also, TGF beta 1 increased apoptosis as measured by M30 labeling at 60 and 180 min (p < 0.001), and by morphological features at 12 h (p < 0.05). In addition, we observed higher levels of activated caspase 3 (17 kDa) from 0 to 30 min. Altogether, these data indicate a direct effect of TGF beta 1 signaling through Smads on both inhibiting proliferation, through alteration of cycle proteins, and inducing apoptosis of gastric epithelial cells in vivo. Further, the studies suggest a potential role for both milk and tissue-expressed TG beta 1 in gastric growth during postnatal development, (C) 2007 Elsevier B.V. All rights reserved.