Ruminal Epithelium Of Sheep Research Articles

Possible influence of free fatty acid receptors on pH regulation in the ruminal epithelium of sheep.

High amounts of short-chain fatty acids (SCFAs) occur in the ovine rumen and constitute the animal's main energy source. However, they lead to an acidification of the ruminal epithelium. Therefore, effective intracellular pH (pHi ) regulation by transport proteins like monocarboxylate transporter 1 (MCT1) and Na+ /H+ exchangers (NHEs) is pivotal to ruminants to avoid epithelial damage. SCFAs might function not only as nutrients but also as signalling molecules by activating free fatty acid receptors (FFARs) in the ruminal epithelium and thus influence pHi regulation. FFARs work as nutrient sensors, transducing their information by modulating cyclic adenosine monophosphate (cAMP) levels. We hypothesized that (FFAR-modulated) decreases in cAMP levels stimulate the activity of MCT1 and NHEs in the ruminal epithelium of sheep. We detected two FFARs (GPR109A and FFAR2) immunohistochemically in the ovine ruminal epithelium. Administration of 10mM butyrate to Ussing chamber-mounted epithelia provoked a significant reduction in intraepithelial cAMP levels. However, application of the GPR109A agonist niacin did not affect cAMP levels. MCT1 activity was analysed by measuring transepithelial 14 C-acetate fluxes, which were not inhibited by forskolin-induced increased cAMP levels. The recovery of pHi after acidification was assessed as an indicator of NHE activity in primary cultured ruminal epithelial cells. Recovery was significantly reduced when cells with increased cAMP levels were subjected to the NHE inhibitor 5-(N-ethyl-N-isopropyl)-amiloride (10µM). Nonetheless, with augmented cAMP levels alone, NHE activity tended to decline. We hypothesize that modulation of cAMP levels by butyrate is accomplished by FFAR2 activation, regulating NHE activity for pHi homoeostasis at least in part.

Permeation of acetate across sheep ruminal epithelium is partly mediated by an anion channel

The present study pursued the question if an anion channel could be partly responsible for the transport of acetate in the isolated ovine ruminal epithelium. Using the Ussing chamber technique, changes of short-circuit current (Isc) induced by mucosal or serosal addition of acetate was investigated. To further evaluate the Isc changes induced by acetate the epithelia were preincubated with ouabain or 4,4′-diisothiocyano-2,2′-stilbenedisulfonic acid (DIDS). In addition, unidirectional flux rates of 14C-acetate were measured at different transepithelial potential difference (Vt). At Vt=0mV, acetate addition to the mucosal side of epithelia incubated in ouabain/DIDS free solution resulted in an Isc decrease, whereas application to the serosal side induced an Isc increase. Absolute Isc changes were significantly larger after serosal than after mucosal acetate addition. Ouabain pre-incubation abolished these side-specific differences. Pre-incubation with DIDS on the mucosal side inhibited the current induced by subsequent mucosal acetate addition in a voltage dependent manner, whereas serosal DIDS pre-incubation had no effect on acetate-induced changes of Isc. Mucosal-to-serosal acetate flux was partly voltage-dependent. The serosal-to-mucosal flux of acetate was not influenced by Vt. The asymmetric changes of Isc after acetate application and the Vt dependence of the DIDS inhibition on the mucosal side indicate a partly electrogenic transcellular permeation of acetate which includes an apically located acetate conductance. It is suggested that this conductance may be important for epithelial cell homeostasis.

Bicarbonate-dependent and bicarbonate-independent mechanisms contribute to nondiffusive uptake of acetate in the ruminal epithelium of sheep

The present study investigated the significance of apical transport proteins for ruminal acetate absorption and their interaction with different anions. In anion competition experiments in the washed reticulorumen, chloride disappearance rate (initial concentration, 28 mM) was inhibited by the presence of a short-chain fatty acid mixture (15 or 30 mM of each acetate, propionate, and butyrate). Disappearance rates of acetate and propionate, but not butyrate (initial concentration, 25 mM each) were diminished by 40 or 80 mM chloride. In isolated ovine ruminal epithelia mounted in Ussing chambers, an increase in chloride concentration from 4.5 to 90 mM led to a decrease of apical acetate uptake at a concentration of 0.5 mM. Mucosal nitrate inhibited acetate uptake most potently whereas sulfate had no effect. Decreasing mucosal pH from 7.4 to 6.1 approximately doubled uptake of acetate both at 0.5 and 10 mM, but this doubling was almost abolished when HCO(3)(-) was absent. The stimulated uptake at mucosal pH 6.1 consisted of a bicarbonate-dependent, nitrate-inhibitable part (K(m) = 54 mM) and a bicarbonate-independent component (K(m) = 12 mM) that was also sensitive to nitrate inhibition. Maximal uptake was three times larger for bicarbonate-dependent vs. bicarbonate-independent uptake. Mucosal addition of 200 microM DIDS, 400 microM p-chloromercuribenzene sulfonic acid, 800 microM p-hydroxymercuribenzoic acid, or 100 microM phloretin had no effects on acetate uptake although the latter two inhibited l-lactate uptake. Our data conclusively show a dominant involvement of proteins in apical acetate uptake. Previously described pH effects on acetate absorption originate mainly from modulation of acetate/bicarbonate exchange. Additionally, there is bicarbonate-independent uptake of acetate anions that is protein coupled but not via monocarboxylate cotransporter.

Apical sodium–glucose co-transport can be regulated by blood-borne glucose in the ruminal epithelium of sheep (Ovis aries, Merino breed)

The intestinal Na-dependent D-glucose co-transporter (SGLT)-1 in sheep is under dietary regulation by luminal substrates. The aim of the present study was to find out whether the SGLT-1 in the forestomach of sheep is also regulated by sugars. Furthermore, the location of a possible glucosensor (luminal v. intracellular v. basolateral) was to be elucidated. Ruminal epithelia of sheep (Ovis aries, Merino breed) were pre-incubated in Ussing chambers with various substrates on the mucosal (i.e. luminal) or serosal (i.e. blood) side. This pre-incubation period was followed by a second pre-incubation period without the tested substrates (washout period). Thereafter, apical D-glucose uptake by ruminal epithelial cells was determined with 200 mumol D-[(14)C]glucose/l in the absence or co-presence of the SGLT-1 inhibitor, phlorizin. Pre-incubation with D-glucose on the mucosal side had no significant effect on apical D-glucose uptake (P>0.05). In contrast, pre-incubation with D-glucose, D-mannose, 3-O-methyl-d-glucose or sucrose on the serosal side significantly increased D-glucose uptake compared with mannitol-treated controls (P<0.05). Serosal pre-incubation with cellobiose or D-xylose had no effect. The stimulation of d-glucose uptake by serosal D-glucose pre-incubation was concentration dependent, with maximal stimulation at about 10 mmol/l. We conclude that the ruminal SGLT-1 can be up-regulated in a concentration-dependent manner by blood-borne D-glucose via an extracellular sugar-sensing mechanism.

Effect of ammonia on Na+ transport across isolated rumen epithelium of sheep is diet dependent.

The cellular uptake of ammonia affects the intracellular pH (pHi) of polar and non-polar cells. A predominant uptake of NH3 and its intracellular protonation tend to alkalinise the cytoplasm, whereas a predominant uptake of NH4(+) acidifies the cytoplasm by reversing this reaction. Hence, the well-known absorption of ammonia across the rumen epithelium probably causes a change in the pHi. The magnitude and direction of this change in pHi (acid or alkaline) depends on the relative transport rates of NH3 and NH4(+). Consequently, the intracellular availability of protons will influence the activity of the Na(+)-H(+) exchanger, which could affect transepithelial Na(+) transport. The aim of the present study has been to test this possible interaction between ruminal ammonia concentrations and Na(+) transport. The term ammonia is used to designate the sum of the protonated (NH4(+)) and unprotonated (NH3) forms. Isolated ruminal epithelium of sheep was investigated by using the Ussing-chamber technique in vitro. The present results indicate that ammonia inhibits Na(+) transport across the rumen epithelium of hay-fed sheep, probably by binding intracellular protons and thus inhibiting Na(+)-H(+) exchange. By contrast, ammonia stimulates Na(+) transport in concentrate-fed and urea-fed sheep, which develop an adaptation mechanism in the form of an increased metabolism of ammonia in the rumen mucosa and/or an increased permeability of rumen epithelium to the charged ammonium ion (NH4(+)). Intracellular dissociation of NH4(+) increases the availability of protons, which stimulate Na(+)-H(+) exchange. This positive effect of ruminal ammonia on Na(+) absorption may significantly contribute to the regulation of osmotic pressure of the ruminal fluid, because intraruminal ammonia concentrations up to 40 mmol/l have been reported.

Transport of ketone bodies and lactate in the sheep ruminal epithelium by monocarboxylate transporter 1.

Due to intensive intracellular metabolism of short-chain fatty acids, ruminal epithelial cells generate large amounts of D-beta-hydroxybutyric acid, acetoacetic acid, and lactic acid. These acids have to be extruded from the cytosol to avoid disturbances of intracellular pH (pH(i)). To evaluate acid extrusion, pH(i) was studied in cultured ruminal epithelial cells of sheep using the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Extracellular addition of D-beta-hydroxybutyrate, acetoacetate, or lactate (20 mM) resulted in intracellular acidification. Vice versa, removing extracellular D-beta-hydroxybutyrate, acetoacetate, or lactate after preincubation with the respective monocarboxylate induced an increase of pH(i). Initial rate of pH(i) decrease as well as of pH(i) recovery was strongly inhibited by pCMBS (400 microM) and phloretin (20 microM). Both cultured cells and intact ruminal epithelium were tested for the possible presence of proton-linked monocarboxylate transporter (MCT) on both the mRNA and protein levels. With the use of RT-PCR, mRNA encoding for MCT1 isoform was demonstrated in cultured ruminal epithelial cells and the ruminal epithelium. Immunostaining with MCT1 antibodies intensively labeled cultured ruminal epithelial cells and cells located in the stratum basale of the ruminal epithelium. In conclusion, our data indicate that MCT1 is expressed in the stratum basale of the ruminal epithelium and may function as a main mechanism for removing ketone bodies and lactate together with H+ from the cytosol into the blood.

Glucose Uptake via SGLT-1 Is Stimulated by β2-Adrenoceptors in the Ruminal Epithelium of Sheep

Glucose absorption via the sodium glucose-linked transporter (SGLT)-1, decreases the glucose concentration in the ruminant forestomach and may ameliorate or prevent ruminal lactic acidosis. Because acidotic ruminants show increased sympathetic activity, the possibility of adrenergic modulation of SGLT-1 was investigated. Glucose uptake into ovine ruminal epithelia was measured in Ussing chambers after the addition of 200 micromol/L (14)C-labeled glucose to the mucosal solution. Glucose uptake decreased (P < 0.05) by >50% in comparison with control after mucosal addition of the SGLT-1 inhibitor, phlorizin (100 micromol/L). Serosal preincubation with 100 micromol/L epinephrine increased (P < 0.05) the phlorizin-sensitive glucose uptake in the absence and presence of indomethacin (10 micromol/L). The effect of epinephrine was simulated by beta- (100 micromol/L isoproterenol) and beta(2)-receptor agonists (10 micromol/L terbutaline), as well as by direct stimulation of adenylyl cyclase (10 micromol/L forskolin). The serosal addition of methoxamine, clonidine, dobutamine or BRL 37344 had no effect. Inhibition of protein kinase A with 2 micromol/L H 89 completely abolished the stimulation of glucose uptake by epinephrine. We conclude that ruminal SGLT-1 can be stimulated via beta(2)-dependent generation of cyclic adenosine monophosphate.

Influence of food deprivation on the transport of 3-O-methyl-alpha-D-glucose across the isolated ruminal epithelium of sheep.

Recent studies provided evidence that the ruminal epithelium is able to absorb D-glucose even at physiologically low intraruminal concentrations. To elucidate whether ruminal D-glucose transport shows adaptive responses during food deprivation, transport of 3-0-methyl-alpha-D-glucose (3-OMG), a hardly metabolizable D-glucose analogue, was measured in isolated ruminal epithelia obtained from hay-fed or food-deprived adult sheep. In both groups, a significant net absorption of 3-OMG to the serosal side (in vivo: blood side oriented) could be detected at 3-OMG concentrations between 0.25 mM and 5 mM. Net absorption of 3-OMG was abolished by mucosal (in vivo: lumen side oriented) addition of phlorizin, an inhibitor of the sodium glucose-linked transporter 1 (SGLT-1). Net absorption of 3-OMG followed Michaelis-Menten kinetics, but apparent affinity and maximal transport capacity were lower in epithelia obtained from food-deprived sheep. In contrast to the decrease of the (secondary) active 3-OMG transport, serosal-to-mucosal permeation of 3-OMG increased after food deprivation, suggesting an elevated passive 3-OMG transfer. It is concluded that the altered transport characteristics are either part of a global energy-sparing process during food deprivation (i.e., a lowered activity of the Na+/K+-ATPase) or result from specific down-regulation of SGLT-1.

Influence of isoform and DNP on butyrate transport across the sheep ruminal epithelium

Short-chain fatty acids are absorbed in considerable amounts from the rumen. During transit through the epithelial layer, they are intensively metabolised. Interaction between intraepithelial metabolism and absorption, however, is hardly understood. The present study therefore compared the transepithelial transport of the easily metabolised n-butyrate with that of the more metabolism-resistant iso-butyrate both under in vivo conditions (isolated and washed reticulorumen) and in vitro conditions (Ussing chamber). Under in vivo conditions, net absorption of n-butyrate was significantly higher than that of iso-butyrate. The in vitro experiments showed that the higher net flux of n-butyrate was solely due to a higher mucosal-to-serosal flux, whereas the serosal-to-mucosal flux of butyrate was independent from the isoform. Blocking intraepithelial ATP delivery by 2,4-dinitrophenol abolished the net flux of n-butyrate. The study indicates that metabolism and/ or ATP availability stimulates n-butyrate net absorption. By this, the metabolic activity of the epithelium may have a regulatory influence on absorption of n-butyrate.

Transport, catabolism and release of histamine in the ruminal epithelium of sheep.

Whereas intraruminal histamine does not affect healthy ruminants, histaminosis is apparent during ruminal acidosis. We therefore investigated the factors that, under physiological circumstances, prevent intoxication by intraruminal histamine and the disturbances occurring during acidotic or hypoxic epithelial damage. After mucosal (m) or serosal (s) application of 80 microM histamine, its flux across the isolated epithelia of the sheep rumen was determined radioactively (hist-rad flux) in Ussing chambers. The non-catabolized component of the hist-rad fluxes was determined by high-pressure liquid chromatography (HPLC) (histamine flux). The difference between hist-rad and histamine fluxes indicated efficient intraepithelial catabolism of histamine at pH 7.4 (m-s direction, 98.7%; s-m direction, 93.3%). Both 0.1 mM 2,4-dinitrophenol (DNP) and mucosal acidification to pH 5.1 increased hist-rad fluxes and decreased catabolic efficiency. pH-dependent secretion of histamine was indicated by differences between m-s and s-m fluxes of histamine and/or hist-rad. Epithelial permeability to hist-rad and mannitol was similar and their fluxes correlated partly. Epithelial release of endogenous histamine was 1.5 pmol x cm(-2) x h(-1) and was not increased by the mast cell stimulator, compound 48/80 (10 ng x ml(-1)). We conclude that histamine absorption across the intact epithelium is efficiently restricted by a low permeability to histamine in combination with catabolic and secretory processes. Especially increases in paracellular permeability and/or inhibition of catabolism enhance histamine absorption.

Transport, catabolism and release of histamine in the ruminal epithelium of sheep

Transport, catabolism and release of histamine in the ruminal epithelium of sheep

Regulatory role of cAMP in transport of Na+, Cl- and short-chain fatty acids across sheep ruminal epithelium.

Sodium is absorbed in considerable amounts across the ruminal epithelium, whilst its transport is strongly interrelated with the permeation of chloride and short-chain fatty acids (SCFAs). However, regulation of ruminal Na+, Cl-, and SCFA absorption is hardly understood. The present study was therefore performed to characterize the influence of cAMP on sodium and sodium-coupled transport mechanisms in short-circuited, stripped ruminal epithelia of sheep. Elevation of intracellular cAMP concentrations by theophylline (10 mM) or theophylline in combination with forskolin (0.1 mM) significantly reduced mucosal-to-serosal sodium transport, leading to a reduction of net transport. The theophylline- or theophylline-forskolin-induced reduction of sodium transport was accompanied by a decrease in chloride net transport but revealed no effect on propionate flux. Short-chain fatty acids stimulated Na+ transport but their stimulatory effect was almost completely blocked by theophylline-forskolin. In solutions with and without SCFAs, the inhibitory effect of 1 mM amiloride on sodium transport was strongly reduced after theophylline-forskolin pretreatment of the tissues. Blocking the production of endogenous prostaglandins by addition of indomethacin (10 microM) led to a theophylline-sensitive stimulation of unidirectional and net fluxes of sodium. The findings indicate that apical, amiloride-sensitive Na+-H+ exchange and/or basolateral Na+-K+-ATPase can effectively be blocked by cAMP, leading to a decrease in sodium and chloride transport. In the ruminal epithelium, cAMP is a second messenger of prostaglandins, which are released spontaneously under in vitro conditions.

Regulatory role of cAMP in transport of Na+, Cl- and short-chain fatty acids across sheep ruminal epithelium

Sodium is absorbed in considerable amounts across the ruminal epithelium, whilst its transport is strongly interrelated with the permeation of chloride and short-chain fatty acids (SCFAs). However, regulation of ruminal Na+, Cl-, and SCFA absorption is hardly understood. The present study was therefore performed to characterize the influence of cAMP on sodium and sodium-coupled transport mechanisms in short-circuited, stripped ruminal epithelia of sheep. Elevation of intracellular cAMP concentrations by theophylline (10 mM) or theophylline in combination with forskolin (0·1 mM) significantly reduced mucosal-to-serosal sodium transport, leading to a reduction of net transport. The theophylline- or theophylline-forskolin-induced reduction of sodium transport was accompanied by a decrease in chloride net transport but revealed no effect on propionate flux. Short-chain fatty acids stimulated Na+ transport but their stimulatory effect was almost completely blocked by theophylline-forskolin. In solutions with and without SCFAs, the inhibitory effect of 1 mM amiloride on sodium transport was strongly reduced after theophylline-forskolin pretreatment of the tissues. Blocking the production of endogenous prostaglandins by addition of indomethacin (10 µM) led to a theophylline-sensitive stimulation of unidirectional and net fluxes of sodium. The findings indicate that apical, amiloride-sensitive Na+-H+ exchange and/or basolateral Na+-K+-ATPase can effectively be blocked by cAMP, leading to a decrease in sodium and chloride transport. In the ruminal epithelium, cAMP is a second messenger of prostaglandins, which are released spontaneously under in vitro conditions.

Tissue distribution of a peptide transporter mRNA in sheep, dairy cows, pigs, and chickens.

A 446-bp cDNA fragment encoding a peptide transport protein was cloned from sheep omasum and used as a probe to study the distribution of the peptide transport protein mRNA in various tissues of sheep, dairy cows, pigs, and chickens. Because the predicted amino acid sequence of this fragment was 85.8, 90.5, and 90.5% identical to rabbit, human, and rat intestinal peptide transporter (PepT1), respectively, it is believed that this cloned fragment represents PepT1 from sheep. In sheep (n = 5) and lactating Holstein cows (n = 3), hybridization was observed with mRNA from the omasum, rumen, duodenum, jejunum, and ileum. The estimated size of mRNA was 2.8 kb. No hybridization was observed with mRNA from the abomasum, cecum, colon, liver, kidney, and semitendinosus and longissimus muscles of either species or the mammary gland of the dairy cows. In pigs (n = 6), the probe hybridized with mRNA from the duodenum, jejunum, and ileum. There was no hybridization with mRNA from the stomach, large intestine, liver, kidney, and semitendinosus and longissimus muscles. Two bands, 3.5 and 2.9 kb, were observed with northern blot analysis, indicating two RNA transcripts that may result from alternative mRNA processing. In White Leghorns (n = 15) and broilers (n = 20), the strongest hybridization was found in the duodenum, but the jejunum and ileum showed faint bands. The size of mRNA in chickens was 1.9 kb. Other tissues, including the crop, proventriculus, gizzard, ceca, liver, kidney, and muscles showed no hybridization to the probe. In conclusion, mRNA for PepT1 is present in the small intestine of all animals examined and the omasal and ruminal epithelium of sheep and dairy cows. The size of the mRNA varied among species.

Identification of two cDNA clones encoding small proline-rich proteins expressed in sheep ruminal epithelium.

Small proline-rich (SPRR) proteins are markers frequently associated with squamous cell differentiation. They have been proposed to be a novel group of precursor polypeptides for the cornified envelope in epidermal keratinocytes. A plus/minus screening procedure was used to identify cDNA clones expressed in mature but not in neonatal sheep ruminal epithelium. Two clones encoding SPRR proteins were identified and are reported here. Clone 27 encodes an ovine SPRR protein corresponding to the human type-II SPRR protein. Clone 26 encodes an ovine SPRR protein similar to human type-II SPRR protein, but which also contains an N-terminal His-Pro repeat similar to the paired repeats found in the Drosophila paired proteins. The unique combination of a paired domain and an SPRR protein has not been reported prior to this study. The tissue distribution indicates that specific expression of the genes corresponding to these two clones occurs in the epithelium of the ruminant forestomach, and to a lesser extent in skin epithelium. In situ hybridization demonstrated that the SPRR mRNA for both clones were localized in the stratum granulosum, in support of their putative physiological function, i.e. formation of the cornified envelope. Based on Northern blot analysis, mRNA complementary to the two clones appears in the ruminal epithelium by 1 week of age, corresponding to the formation of the stratum granulosum during ruminal epithelial development. The different patterns of changes in amount of mRNA corresponding to these clones during rumen epithelial development indicate that they play different roles in rumen epithelial development.

Absorption of short-chain fatty acids across ruminal epithelium of sheep.

Investigations on the absorption of shortchain fatty acids across ruminal epithelium of sheep were performed both in vitro (Ussing chamber technique, using propionic acid representatively for short-chain fatty acids) and in vivo (washed, isolated reticulorumen). A pH-induced, nearly tenfold increase in the concentration of undissociated propionate led to an only twofold increase in mucosal-to-serosal flux of propionate (in vitro). Neither amiloride (1 mmol.l-1, in vitro) nor theophylline (10 mmol.l-1, in vivo), inhibitors of the ruminal Na+/H+ exchanger, exerted any significant influence on propionate fluxes or short-chain fatty acids absorption, respectively. Total replacement of luminal Na+ (by choline) did not alter short-chain fatty acids absorption (in vivo). Mucosal 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (0.1 mmol.l-1) or mucosal nitrate (40 mmol.l-1) markedly reduced propionate net flux (in vitro). Increasing mucosal Cl- concentration brought about a significant drop in mucosal-to-serosal flux of propionate (in vitro) and in short-chain fatty acids net absorption (in vivo), respectively. The results obtained suggest that short-chain fatty acids are absorbed both as anions and as undissociated acids across ruminal epithelium of sheep. It is concluded that short-chain fatty acids anions either compete with Cl- for binding sites at a common anion-exchange mechanism or that they are absorbed by an short-chain fatty acids anion/ HCO3- exchanger indirectly coupled to a Cl-/HCO3- exchanger via intracellular bicarbonate.

Isolation and characterization of a cDNA clone encoding ovine type I carbonic anhydrase.

To identify genes involved in the postnatal development of sheep ruminal epithelium, a lambda gt22a cDNA library was constructed from poly(A)+ RNA isolated from mature sheep ruminal epithelium. A plus/minus screening procedure was used to identify genes expressed in mature but not in neonatal ruminal epithelium. One of the cDNA clones isolated encodes an ovine carbonic anhydrase, based on nucleotide and deduced peptide sequence analysis. The deduced peptide is most closely related to eukaryotic type I carbonic anhydrase, based on comparison with all available carbonic anhydrase sequences. Northern blot hybridization confirmed that the amount of mRNA complementary to the carbonic anhydrase cDNA clone is more than five times higher in ruminal epithelium for mature sheep (12 wk old) than in ruminal epithelium from neonatal lambs (< 12h old). Messenger RNA complementary to this cDNA clone was found only in the epithelium of the ruminant forestomach compartments (i.e., rumen, reticulum, and omasum), but small amounts of hybridizable mRNA were also found in sheep skin. This carbonic anhydrase cDNA clone will allow the study of transcriptional regulation of the carbonic anhydrase gene during ruminal epithelial development.

Developmental Changes in Glucose and Butyrate Metabolism by Isolated Sheep Ruminal Cells ,

The ontogeny of glucose and butyrate metabolism in developing sheep ruminal epithelium was determined using an isolated ruminal cell system. Ruminal cells were isolated from 21 lambs at seven ages before weaning. Rumen weight increased in proportion to increases in body weight, except between 28 and 42 d, when rumen weight increased threefold, whereas body weight increased only 33%. Glucose oxidation rates [expressed as nmol/(1 × 106 ruminal cells·120 min)] by ruminal cells were low at birth (14.2 ± 5.08), increased sharply by 14 d (71.38 ± 16.71), and remained elevated until 42 d. Following 42 d, glucose oxidation declined to rates lower than those observed at birth (6.11 ± 0.83). Butyrate oxidation to CO2 increased from birth (20.03 ± 3.41) to a peak at 4 d (134.0 ± 31.71) and decreased throughout the remainder of the preweaning period (56 d; 36.32 ± 7.48). Butyrate inhibited glucose oxidation by ruminal cells isolated at 14, 28 and 42 d. Similarly, glucose inhibited butyrate oxidation by ruminal cells isolated from 4 d to 28 d following birth. β-Hydroxybutyrate production [nmol/(1 × 106 ruminal cells·120 min)] from butyrate by ruminal cells was undetectable at birth, but measurable by 4 d (3.28 ± 2.15). Rates of β-hydroxybutyrate production remained unchanged through 42 d; however, by 56 d, production had increased 10-fold (36.71 ± 0.67). The metabolic adaptations of the ruminal epithelium are intimately associated with the physical development of the tissue, and major shifts in the fate of glucose and butyrate carbon occur prior to weaning.

Water and sodium movements across the ruminal epithelium in fed and food‐deprived sheep

Fluid and electrolyte movements across the ruminal epithelium of sheep were studied using the temporarily isolated rumen technique. The sheep were all subjected to the following treatments: (1) fed sheep (fed twice daily), after rumen emptying, received rumen contents from a fed sheep; (2) food-deprived sheep (two meals were omitted), after rumen emptying, received rumen contents from a food-deprived sheep; (3) fed sheep, after rumen emptying, received rumen contents from a food-deprived sheep; and (4) food-deprived sheep, after rumen emptying, received rumen contents from a fed sheep. Food deprivation led to an increased Na concentration of the rumen fluid while K and Cl concentrations, as well as osmolality, decreased. Plasma Na and osmolality decreased. During the 40 min after the rumen contents were exchanged no net movements of water occurred. Then the sheep were given an intraruminal load of saline which gave rise to a significant net absorption of fluid from the rumens of those sheep which had received rumen contents from fed sheep. The change in composition of the rumen contents after food deprivation impaired the absorption of Na and water from the rumen. Furthermore food deprivation reduced the Na absorptive function of the ruminal epithelium, but not the water permeability.

The occurrence of paranuclear vacuoles in sheep ruminal epithelium treated with distilled water in vitro and from sheep injected with a large quantity of water into the rumen

In the histological preparation taken after animal death, paranuclear vacuoles (PV) in sheep ruminal epithelium were already present in high percentage (7.5-20.3%). With the incubation of the ruminal mucosae in distilled water (10-15 degrees C) up to 120 min, PV occurrence did not alter. With incubation in NaCl solution, however, PV value decreased with time proportional to NaCl concentration (0.5, 0.85 or 1.5%). In the preparation taken by biopsy, on the other hand, PV were rare in ruminal epithelium (less than 0.3%). With the injection of warm water (10-141/animal), however, PV occurrence in the ruminal epithelium was not affected. PV as cytoplasmic processes of the ruminal Langerhans cells are probably formed by the reaction of these cells to environmental changes in the tissue caused by animal death.