Small Intestine Of Suckling Rats Research Articles

Effect of Misoprostol on the enzyme ontogeny of the rat intestine

The effect of Misoprostol (an analog of PGE1) on the biochemical changes in the small intestine of suckling rats was studied. Misoprostol increases sucrase activities in the proximal and the distal small intestine. Jejunal aminopeptidase N activity is higher in Misoprostol-treated rats than in the control rats. This drug also modifies the relative weight of the small intestine and the mucosal ratio of DNA to RNA. Misoprostol effects appear to be mediated by corticosterone release.

Effect of Misoprostol on the enzyme ontogeny of the rat intestine

The effect of Misoprostol (an analog of PGE1) on the biochemical changes in the small intestine of suckling rats was studied. Misoprostol increases sucrase activities in the proximal and the distal small intestine. Jejunal aminopeptidase N activity is higher in Misoprostol-treated rats than in the control rats. This drug also modifies the relative weight of the small intestine and the mucosal ratio of DNA to RNA. Misoprostol effects appear to be mediated by corticosterone release.

The Presence of Transforming Growth Factor-α in the Suckling Rat Small Intestine and Pancreas and the Absence in Rat Milk

Immunoreactive rat transforming growth factor-alpha (TGF-alpha) was measured in rat milk, in the mucosa and lumen of the small intestine, and in the pancreas of suckling and adult rats with a homologous RIA. In contrast to epidermal growth factor, where the main source of epidermal growth factor for sucklings is rat milk, the presence of TGF-alpha was not detectable in rat milk. The concentrations of TGF-alpha in the small intestine exhibited similar values in suckling and adult rats, whereas epidermal growth factor levels in the small intestine were several times higher in suckling rats than adults. Overnight fasting in suckling rats resulted in minimal changes in the luminal and mucosal TGF-alpha content. A positive correlation was established in suckling rats between the TGF-alpha content in the intestinal lumen (but not mucosa) and the TGF-alpha content in pancreas. Despite many structural and functional similarities between TGF-alpha and epidermal growth factor, our present data indicate significant differences in the origin and distribution of these two growth factors in the tract of the small intestine of developing rats.

Remodeling of the Rat Small Intestinal Mucosa During the Suckling Period

The development of the suckling rat small intestine has been reexamined to investigate changes to cell kinetics and morphology more fully. In this period of rapid growth and reorganization, villi and crypts steadily increase in size in proximal regions; however, in distal regions a dramatic shortening of villi occurs at about 15 days. The result of this remodeling is that the normal adult proximodistal gradient of villus height is established. At 15 days, migration rates in both regions increase dramatically. In proximal regions, the extent of 24‐h migration of thymidine‐labeled enterocytes is dependent on villus height. In distal regions, the increase in migration rate can be explained by the shortening of the villi. The extent of 24‐h migration was not related to villus height. On the basis of these results, it is suggested that care must be taken in drawing conclusions relating any parameter to assumed distal region cell renewal patterns. This is because changes in the relative proportions of cell types present apparently can be brought about by villus deletion, independent of cell renewal.

Control of fatty acid oxidation by intramitochondrial [NADH]/[NAD+] in developing rat small intestine.

The oxidation of palmityl-coenzyme A and acetate to CO2 by mitochondria isolated from rat small intestine increases 10-fold at the time of weaning (18-21 days of age). Carnitine palmitoyltransferase (EC 2.3.1.21) activity is 2-fold greater in mitochondria of suckling rat intestine compared to postweaned intestine. These data indicate that carnitine palmitoyltransferase does not control the increase in intestinal fatty acid oxidation during weaning. We have previously reported that the estimated intramitochondrial [NADH]/[NAD+] as determined by the ratio of tissue levels of 3-hydroxybutyrate and acetoacetate is fivefold greater in suckling rat intestine compared to postwean animals. High intramitochondrial [NADH]/[NAD+] which is present in suckling rat small intestine is associated with a decrease in citric acid cycle activity and beta oxidation. The addition of acetoacetate causes a decrease in intramitochondrial [NADH]/[NAD+]. The oxidation of acetate and glucose to CO2 by suckling rat intestine mitochondria was stimulated by the addition of 1 mM acetoacetate. These data suggest that the lower rate of fatty acid oxidation by suckling rat small intestine is controlled by elevated intramitochondrial [NADH]/[NAD+].

Precocious Increase of Sucrase Activity by Carbohydrates in the Small Intestine of Suckling Rats. II. Role of Digestibility of Sugars, Osmolality, and Stomach Evacuation in Producing Diarrhea

SummaryThe mechanisms of carbohydrate induced diarrhea in suckling rats were investigated with respect to osmolality and type of sugar in the milk. Groups of 12 day old rats were gavage fed either a basic low carbohydrate milk formula [10.8% fat; 8% protein; 1.4% carbohydrate (weight/volume)] or basic formula with added sucrose, fructose, lactose, or glucose polymers, all as 13% (weight/ volume). All formulas were isocaloric. Their corresponding osmolalities were 278, 645, 1,130, 617, and 349 mOsmol/ kg, respectively. Gastric evacuation of water soluble materials from formulas containing sucrose, fructose, or glucose polymers was significantly slower than the gastric evacuation of the basic formula and the formula that contained lactose. The net fluid absorption from the small intestine was significantly greater from the basic and lactose containing formulas when compared with sucrose, fructose, or glucose polymer containing formulas. When the synthetic milk formulas were placed directly into the isolated noileum in vivo, the formulas of higher osmolality (fructose, sucrose, and lactose) caused water flux into the intestine at 60 min, while digestion of the lactose formula reversed the water flux within 120 min. We conclude that the type of added sugar is a decisive factor in gastric evacuation, and that water flux into and out of the intestine is significantly affected by the osmolality and rate of digestion and absorption of the carbohydrate in the formula; these differences among sugars may play a significant role in the etiology of diarrhea.

Precocious increase of sucrase activity by carbohydrates in the small intestine of suckling rats. II. Role of digestibility of sugars, osmolality, and stomach evacuation in producing diarrhea.

The mechanisms of carbohydrate-induced diarrhea in suckling rats were investigated with respect to osmolality and type of sugar in the milk. Groups of 12-day-old rats were gavage fed either a basic low carbohydrate milk formula [10.8% fat; 8% protein; 1.4% carbohydrate (weight/volume)] or basic formula with added sucrose, fructose, lactose, or glucose polymers, all as 13% (weight/ volume). All formulas were isocaloric. Their corresponding osmolalities were 278, 645, 1,130, 617, and 349 mOsmol/kg, respectively. Gastric evacuation of water-soluble materials from formulas containing sucrose, fructose, or glucose polymers was significantly slower than the gastric evacuation of the basic formula and the formula that contained lactose. The net fluid absorption from the small intestine was significantly greater from the basic and lactose-containing formulas when compared with sucrose, fructose, or glucose polymer-containing formulas. When the synthetic milk formulas were placed directly into the isolated jejunoileum in vivo, the formulas of higher osmolality (fructose, sucrose, and lactose) caused water flux into the intestine at 60 min, while digestion of the lactose formula reversed the water flux within 120 min. We conclude that the type of added sugar is a decisive factor in gastric evacuation, and that water flux into and out of the intestine is significantly affected by the osmolality and rate of digestion and absorption of the carbohydrate in the formula; these differences among sugars may play a significant role in the etiology of diarrhea.

Precocious increase of sucrase activity by carbohydrates in the small intestine of suckling rats. I. Significance of the stress effect of sugar-induced diarrhea.

In this paper, we analyze the factors involved in the precocious increase of sucrase activity evoked by the early feeding of sucrose in suckling rats, and particularly, the role of diarrhea and stress in this phenomenon. Ten-day-old rats were removed from their mothers and gavage fed for 4 days at 3-h intervals either a basic low carbohydrate milk formula (10.8% fat, 8% protein, 1.4% carbohydrate; all by weight/volume) or basic low carbohydrate milk with: lactose (13%), fructose (13%), or Polycose (2%, 6%, or 13%); all formulas were isocaloric. Feeding the formula containing fructose or high (13%) Polycose led to diarrhea and evoked a concurrent increase of small intestinal sucrase activity. In further experiments, 11-day-old rats were fed the basic formula, the lactose (13%), the fructose (13%), and a sucrose (13%) formula for 8 h between 2 a.m. and 10 a.m. Also, 10-day-old rats were fed 0.5 ml of a solution of 5% mannitol in water while nursing with their mothers. The serum corticosterone levels were substantially increased within 8 h after the initiation of feedings with sucrose and fructose milks and the mannitol solution. The mannitol-fed rats also developed diarrhea within a day in association with a marked increase in sucrase activity. We conclude that a precocious increase of sucrase activity in the small intestine of suckling rats by dietary sugars is not caused by substrate induction, but is mainly due to the effect of stress. The stress is caused by diarrhea which is evoked by the feeding of indigestible and/or unabsorbable amounts of sugar.

Precocious Increase of Sucrase Activity by Carbohydrates in the Small Intestine of Suckling Rats. I. Significance of the Stress Effect of Sugar‐Induced Diarrhea

In this paper, we analyze the factors involved in the precocious increase of sucrase activity evoked by the early feeding of sucrose in suckling rats, and particularly, the role of diarrhea and stress in this phenomenon. Ten‐day‐old rats were removed from their mothers and gavage fed for 4 days at 3‐h intervals either a basic low carbohydrate milk formula (10.8% fat, 8% protein, 1.4% carbohydrate; all by weight/volume) or basic low carbohydrate milk with: lactose (13%), fructose (13%), or Polycose (2%, 6%, or 13%); all formulas were isocaloric. Feeding the formula containing fructose or high (13%) Polycose led to diarrhea and evoked a concurrent increase of small intestinal sucrase activity. In further experiments, 11‐day‐old rats were fed the basic formula, the lactose (13%), the fructose (13%), and a sucrose (13%) formula for 8 h between 2 a.m. and 10 a.m. Also, 10‐day‐old rats were fed 0.5 ml of a solution of 5% mannitol in water while nursing with their mothers. The serum corticosterone levels were substantially increased within 8 h after the initiation of feedings with sucrose and fructose milks and the mannitol solution. The mannitol‐fed rats also developed diarrhea within a day in association with a marked increase in sucrase activity. We conclude that a precocious increase of sucrase activity in the small intestine of suckling rats by dietary sugars is not caused by substrate induction, but is mainly due to the effect of stress. The stress is caused by diarrhea which is evoked by the feeding of indigestible and/or un‐absorbable amounts of sugar.

FATTY ACID OXIDATION BY DEVELOPING RAT INTESTINE IS CONTROLLED BY CHANGES IN MITOCHONDRIAL [NADH]/[NAD

At the time of weaning there is an eight-fold increase in palmitate oxidation to CO2 by rat intestinal mitochondria. Oxidation of acetyl-CoA also increases indicating that the tricarboxyylic acid (TCA) cycle may be rate-limiting during the suckling period. An increase in mitochondrial (mito) [NADH]/[NAD] inhibits isocitrate dehydrogenase, a control enzyme in the TCA cycle. In the suckling rat mito [NADH]/[NAD] is five-fold greater than postwean ratios. If the increase in mito [NADH]/[NAD] during the suckling period regulates TCA cycle activity, decreasing the mito [NADH]/[NAD] should increase TCA cycle activity and fatty acid oxidation. We have determined that high concentrations of potassium inhibit ketone oxidation which results in a decrease in mito [NADH/NAD] by altering the equilibrium of β-hydroxybutyrate dehydrogenase. The oxidation of palmitate and acetate by mitochondria of suckling rat small intestine is activated by the addition of high concentrations of potassium (100 mM) to the mitochondrial isolation mixture. These data suggest that the fatty acid oxidative mechanism is present during the suckling period and an increased mito [NADH]/[NAD] during this period decreases TCA cycle activity and fatty acid oxidation.

Intestinal cathepsin B and D activities of suckling rats.

Cathepsin B and D activities were assayed in homogenates of the mucosa of the proximal, middle, and distal sections of the small intestine of suckling rats of various ages and of adult rats. Cathepsin B activity was determined using N-benzoyl-L-phenylalanyl-L-valyl-L-arginine-p-nitroanilide as substrate in the presence of trypsin inhibitor (pH optimum: 5.0-5.5). Cathepsin D activity was determined by measuring the pepstatin A sensitive action on denatured haemoglobin (pH optimum: 3.0-3.5). In the distal intestine, the activities of both cathepsins increased 6- to 10-fold during the first 2 weeks post partum and then sharply declined to the low values seen in adult rats, but in the proximal intestine the activities were low throughout the suckling period. In vitro digestion of casein by ileal mucosa of 8-day-old rats was almost completely inhibited by the combined actions of leupeptin and pepstatin A. The results show that cathepsin B- and D-like proteinases could play an important role in the digestion of milk proteins by suckling rats.

P-402) - Changeover in alkaline phosphatase of suckling rat small intestine induced by injection of hydrocortisone

P-402) - Changeover in alkaline phosphatase of suckling rat small intestine induced by injection of hydrocortisone

Developmental Transition of Alkaline Phosphatase from Suckling to Adult Type in Rat Small Intestine: Molecular Species and Effect of Injected Cortisone and Thyroxine<xref ref-type="fn" rid="fn1"><sup>1</sup></xref>

Postnatal change of rat intestinal alkaline phosphatase from suckling to adult type occurred at the weaning stage (Uezato et al. (1981) Biochem. Int. 2, 561). Two distinct forms were observed in suckling rat small intestine. One was soluble and the other was membrane-bound. The ratio of the soluble to the membrane-bound form increased after birth until these forms were replaced by the adult type, which consisted of membrane-bound forms different from that of sucklings. The soluble form was demonstrated almost exclusively during the suckling period and was distributed in the distal third of the small intestine. The postnatal increase of alkaline phosphatase was enhanced by administration of hormones such as cortisone and thyroxine. On the other hand, acid β-galactosidase activity in the distal segment was reduced rapidly by the same treatment. The change of alkaline phosphatase from suckling to adult type was accelerated by the injected hormones. In the distal segment the ratio of soluble to membrane-bound activity in the homogenate prematurely decreased after administration of the hormones. Between 16 and 18 days of age, alkaline phosphatase of hormone-treated rat small intestine changed completely from fetal-suckling type to typical adult type in both the proximal and distal segments.

Free amino acid concentrations in the gut lumen of developing rats

Free amino acid concentrations in the lumen and tissue of small intestine of suckling rats were measured by a micromethod. Almost all amino acid concentrations (Lys excepted) in the intestinal juice decrease until 15 days of age whereby the pattern remains constant. Concomitantly, free amino acid concentrations in the intestinal tissue increase between 10 and 15 days p.p. The pool of amino acids in the intestinal juice, reflecting the nutritional input and its utilization, becomes exhausted during the suckling period. At 20 days of age the rats additionally ate the rat chow and free amino acid concentrations in the intestinal tissue increased again. The decline of the amino acid pool in the lumen of small intestine which is both derived by the mother and exhausted by the growing organism may act as a signal for differentiation via (self-) restriction of substrates for protein accretion.

Ultrastructural localization of calcium, byelectron-probe X-ray microanalysis, in the small intestine of suckling rats

The subcellular distribution of calcium has been investigated in samples, from the intestinal mucosa of 10-day-old rats, prepared for X-ray microanalysis by various techniques designed to minimize the loss of this element. Calcium retention and its threshold of detection was most satisfactory in freeze-dried frozen thin sections. In resin-embedded samples the best retention of calcium was found in specimens fixed in absolute ethanol, embedded without osmication, and sectioned onto glycerol. The results of this investigation indicate the presence of calcium in the supranuclear vacuole of enterocytes in the distal intestine of the neonatal rat. This calcium is probably taken up during the endocytosis of material from the intestinal lumen. The same mechanism may also be important in the uptake of other metals by suckling animals.

Phosphoenolpyruvate Carboxykinase in the Small Intestine of Developing Rodents

Phosphoenolpyruvate carboxykinase activity was found to be high in the mucosa of the small intestine of suckling rats. Activity was higher proximally than distally and decreased to very low values at weaning. After weaning, feeding a high fat diet or starvation did not result in elevation of the low intestinal activity in rats. However, in mice, who also showed high activity in the suckling period, starvation, but not diet, did cause a rise. Since fructose-diphosphatase was also found in the intestinal mucosa of suckling rats, it is suggested that gluconeogenesis may occur in this tissue in the neonatal period.gluconecogenesis phosphoenolpyruvate carboxykinase small intestine

Isolation and characterization of the proximal and distal forms of lactase-phlorizin hydrolase from the small intestine of the suckling rat

The complex between lactase (β- d-galactoside galactohydrolase, EC 3.2.1.23) and phlorizin hydrolase ( glycosyl-N- acylsphingosine glycohydrolase, EC 3.2.1.62) has been purified from the proximal and distal regions of the small intestine of suckling rats. The two enzymes behaved differently on DEAE-cellulose ion-exchange chromatography and during electrophoresis in the presence and absence of sodium dodecyl sulphate (SDS), but they had very similar cyanoge bromide cleavage patterns. Kinetic studies on the proximal and distal enzymes showed the sama pH optimum of 6.0 and the same heat stability at 45°C, but a small difference in K m . Treatment of both enzymes with fucosidase, mannosidase or N- acetylhexosaminidase did not affect enzymic activity or electrophoretic mobility. Neuraminidase digestion abolished the electrophoretic differences and gave two active enzymes with similar isoelectric points.

Cell kinetics in the small intestine of suckling rats. I. Influence of hypophysectomy.

Cell kinetics have been examined in the duodena of intact rats at 6, 16, and 22 days of age, and of hypophysectomized rats at 22 days. In intact rats the crypt population per villus increases more than 10-fold, the greater part of the increase occurring late in the third week. The labelling index does not differ between 6 (28.3%) and 16 (27.6%) days, but increases to 37.8% at 22 days. Generation time also does not differ between 6 (18.0 hours) and 16 (17.7 hours) days, but falls to 10.8 hours at 22 days by shortening of both the presynthetic and synthetic phases. Acceleration of cell migration rate between 18 and 19 days of age results in shortening of cell transit time from seven days in rats younger than 18 days to two days in those older than 18 days. When rats are hypophysectomized at six days, the duodenal crypt population per villus at 22 days is comparable to that of intact rats at 16 days. The labelling index at 22 days is 19% below that of intact rats at six days. Generation time at 22 days is slightly shortened by a decrease of the presynthetic phase, but the duration of 15.5 hours is 43% longer than in intact rats at the same age. Despite the small crypt population and the low cell production rate, the cell-transient time in 22-day-old hypophysectomized animals is only three days.

POSTNATAL CHANGES OF ACTIVITY OF ACID LIPASE AND NONSPECIFIC NEUTRAL ESTERASE IN RAT LIVER

Hydrolysis of the 4-methylumbelliferyl (4-MU) esters, oleate and nonanoate, was studied in homogenates of liver from suckling and adult rats. In both age groups, the pH optimum for 4-MU oleate hydrolysis was 4.0, whereas 4-MU nonanoate was hydrolyzed maximally at pH 6.5-7.0. Acid lipaee (4-MU oleate, pH 4.0) was resistant to inhibition by the organophosphate, dlethyl-p-nitrophenyl phosphate (E600), while nonspecific neutral esterase (4-MU nonanoate, pH 7.0) was sensitive to inhibition. Acid lipase and nonspecific neutral esterase differed substantially in the patterns of their postnatal development in the liver. Acid lipase activity was high during the suckling period and decreased after the third week of life to adult levels which were 50% of suckling levels. Nonspecific neutral esterase increased markedly with age, and adults attained levels 10 times those of sucklings. These experiments together with our concomitant findings of high acid lipase in the small intestine of suckling rats suggest the important metabolic role of acid lipase during the suckling period when fat intake is high. (Supported in part by NIH grants HD 08536 and HL 18723-01).

Acid hydrolases in the suckling rat small intestine. II. On the importance of alkaline phosphatase inhibition in the histochemical localization of acid phosphatase activity

Phosphatase activities against beta-glycerophosphate, I-naphthyl phosphate and naphthol AS-TR phosphate were investigated, at acid and aldaline pH levels, using unfixed and fixed cryostat sections of suckling rat jejunum. The use of 10 mm EDTA and 10 mm NaF as inhibitors indicated that alkaline phosphate is predominantly located in the microvillous region of the adsorptive cells, while acid phosphatase is located in small particles distributed between the brush borders and the nuclei of these cells. Alkaline phosphatase activity was found to interfere with the localization of acid phosphatase unless EDTA was included in the incubation medium. A modified Gomori medium, containing 10 mm EDTA and additional lead nitrate, is described. Latency experiemtns using this medium, with unfixed sections, indicated the lysosomal nature of particulate acid phosphatase. The discussion stresses the importance of including an aldaline phosphatase inhibitor in incubation media designed to localize extralysosomal acid phosphatase activity.