Highly fermentable diets are rapidly converted to organic acids [i.e., short-chain fatty acids (SCFA) and lactic acid] within the rumen. The resulting release of protons can constitute a challenge to the ruminal ecosystem and animal health. Health disturbances, resulting from acidogenic diets, are classified as subacute and acute acidosis based on the degree of ruminal pH depression. Although increased acid production is a nutritionally desired effect of increased concentrate feeding, the accumulation of protons in the rumen is not. Consequently, mechanisms of proton removal and their quantitative importance are of major interest. Saliva buffers (i.e., bicarbonate, phosphate) have long been identified as important mechanisms for ruminal proton removal. An even larger proportion of protons appears to be removed from the rumen by SCFA absorption across the ruminal epithelium, making efficiency of SCFA absorption a key determinant for the individual susceptibility to subacute ruminal acidosis. Proceeding initially from a model of exclusively diffusional absorption of fermentation acids, several protein-dependent mechanisms have been discovered over the last 2 decades. Although the molecular identity of these proteins is mostly uncertain, apical acetate absorption is mediated, to a major degree, via acetate-bicarbonate exchange in addition to another nitrate-sensitive, bicarbonate-independent transport mechanism and lipophilic diffusion. Propionate and butyrate also show partially bicarbonate-dependent transport modes. Basolateral efflux of SCFA and their metabolites has to be mediated primarily by proteins and probably involves the monocarboxylate transporter (MCT1) and anion channels. Although the ruminal epithelium removes a large fraction of protons from the rumen, it also recycles protons to the rumen via apical sodium-proton exchanger, NHE. The latter is stimulated by ruminal SCFA absorption and salivary Na(+) secretion and protects epithelial integrity. Finally, SCFA absorption also accelerates urea transport into the rumen, which via ammonium recycling, may remove protons from rumen to the blood. Ammonium absorption into the blood is also stimulated by luminal SCFA. It is suggested that the interacting transport processes for SCFA, urea, and ammonia represent evolutionary adaptations of ruminants to actively coordinate energy fermentation, protein assimilation, and pH regulation in the rumen.

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RUMINANT NUTRITION SYMPOSIUM: Role of fermentation acid absorption in the regulation of ruminal pH

Alterations in rumen epithelial structure and function during grain-induced subacute ruminal acidosis (SARA) are largely undescribed. In this study, four mature nonlactating dairy cattle were transitioned from a high-forage diet (HF; 0% grain) to a high-grain diet (HG; 65% grain). After feeding the HG diet for 3 wk, the cattle were transitioned back to the original HF diet, which was fed for an additional 3 wk. Continuous ruminal pH was measured on a weekly basis, and rumen papillae were biopsied during the baseline and at the first and final week of each diet. The mean, minimum, and maximum daily ruminal pH were depressed (P < 0.01) in the HG period compared with the HF period. During the HG period, SARA was diagnosed only during week 1, indicating ruminal adaptation to the HG diet. Microscopic examination of the papillae revealed a reduction (P < 0.01) in the stratum basale, spinosum, and granulosum layers, as well as total depth of the epithelium during the HG period. The highest (P < 0.05) papillae lesion scores were noted during week 1 when SARA occurred. Biopsied papillae exhibited a decline in cellular junctions, extensive sloughing of the stratum corneum, and the appearance of undifferentiated cells near the stratum corneum. Differential mRNA expression of candidate genes, including desmoglein 1 and IGF binding proteins 3, 5, and 6, was detected between diets using qRT-PCR. These results suggest that the structural integrity of the rumen epithelium is compromised during grain feeding and is associated with the differential expression of genes involved in epithelial growth and structure.

Bovine rumen epithelium undergoes rapid structural adaptations during grain-induced subacute ruminal acidosis

Feeding highly fermentable diets to ruminants is one strategy to increase energy intake. The increase in short-chain fatty acid (SCFA) production and reduced ruminal pH associated with highly fermentable diets imposes a challenge to the metabolism and the regulation of intracellular pH homeostasis of ruminal epithelia. The ruminal epithelia respond to these challenges in a coordinated manner. Whereas the enlargement of absorptive surface area is well documented, emerging evidence at the mRNA and transporter and enzyme activity levels indicate that changes in epithelial cell function may be the initial response. It is not surprising that gene expression analysis has identified pathways involved in fatty acid metabolism, ion transport, and intracellular homeostasis to be the pathways dominantly affected during adaptation and after adaptation to a highly fermentable diet. These findings are important because the intraepithelial metabolism of SCFA, particularly butyrate, helps to maintain the concentration gradient between the cytosol and lumen, thereby facilitating absorption. Butyrate metabolism also controls the intracellular availability of butyrate, which is widely regarded as a signaling molecule. Current data indicate that for butyrate metabolism, 3-hydroxy-3-methylglutaryl-CoA synthase and acetyl-CoA acetyltransferase are potential regulatory points with transient up- and downregulation during diet adaptation. In addition to nutrient transport and utilization, genes involved in the maintenance of cellular tight junction integrity and induction of inflammation have been identified as differentially expressed genes during adaptation to highly fermentable diets. This may have important implications on ruminal epithelial barrier function and the inflammatory response often associated with subacute ruminal acidosis. The objective of this review is to summarize ruminal epithelial adaptation to highly fermentable diets focusing on the changes at the enzyme and transporter activity levels, as well as the underlying molecular changes at the mRNA and protein expression levels.

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Ruminant Nutrition Symposium: Molecular adaptation of ruminal epithelia to highly fermentable diets.

We tested the hypothesis that the dietary energy-dependent alterations of the rumen papillae size are accompanied by corresponding changes in systemic insulin-like growth factor (IGF)-1 concentration and in rumen papillary IGF type 1 receptors (IGF-1R). Young male goats (n = 24) were randomly allocated to two groups (n = 12) and fed a high level (HL) metabolizable energy [1200 kJ/(kg 0.75 . d)] or a low level (LL) [500 kJ/(kg 0.75 . d)] diet for 42 d. The concentration of ruminal total SCFA did not differ between the groups, but the molar proportion of butyric acid was enhanced by 70% in the HL group (P < 0.05). Both the length and width of the papillae were greater (P < 0.05) in the HL group, and the surface was 50-100% larger (P < 0.05) in the tissue sampled from the artrium ruminis, the ventral ruminal sac and the ventral blind sac. Transport of Na + across the rumen epithelium, which is amiloride sensitive, was higher (P < 0.05) in the HL than in the LL group. Furthermore, the plasma IGF-1 concentration was about twofold higher in the HL group (P < 0.05), and the maximal rumen epithelial IGF-1 R binding was also higher in the HL (P < 0.05) than in the LL group. IGF-1R mRNA and IGF-1 mRNA were detected in rumen papillae; however, they were unaffected by dietary treatments. DNA synthesis and cell proliferation of cultured rumen epithelial cells were higher (P < 0.05) after IGF-1 treatment (25 or 50 μg/L) compared with those in the medium without IGF-1. Thus dietary energy-dependent alterations of rumen morphology and function are accompanied by corresponding changes in systemic IGF-1 and ruminal IGF-1R.

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An energy-rich diet causes rumen papillae proliferation associated with more IGF type 1 receptors and increased plasma IGF-1 concentrations in young goats.

The ruminal epithelium has an enormous capacity for the absorption of short-chain fatty acids (SCFAs). This not only delivers metabolic energy to the animal but is also an essential regulatory mechanism that stabilizes the intraruminal milieu. The epithelium itself, however, is endangered by the influx of SCFAs because the intracellular pH (pHi) may drop to a lethal level. To prevent severe cytosolic acidosis, the ruminal epithelium is able to extrude (or buffer) protons by various mechanisms: (i) a Na+/H+ exchanger, (ii) a bicarbonate importing system and (iii) an H+/monocarboxylate cotransporter (MCT). Besides pHi regulation, the MCT also provides the animal with ketone bodies derived from the intraepithelial breakdown of SCFAs. Ketone bodies, in turn, can serve as an energy source for extrahepatic tissues. In addition to SCFA uptake, glucose absorption has recently been identified as a potential way of eliminating acidogenic substrates from the rumen. At least with respect to SCFAs, absorption rates can be elevated when adapting animals to energy-rich diets. Although they are very effective under physiological conditions, the absorptive and regulatory mechanisms of the ruminal epithelium also have their limits. An increased number of protons during the state of ruminal acidosis can be eliminated neither from the lumen nor the cytosol, thus worsening dysfermentation and finally leading to functional and morphological alterations of the epithelial lining.

Transfer of energy substrates across the ruminal epithelium: implications and limitations.

Six mature sheep fitted with rumen fistulae underwent four consecutive feeding periods, of 15 weeks each, i.e. (1) hay only; (2) 36% hay, 64% concentrate; (3) 10% hay, 90% concentrate; and (4) hay only again. The net absorption of sodium, chloride and magnesium from the washed rumen filled with artificial rumen fluids was tested during each feeding period. The artificial rumen fluids varied either in osmolarity (315 and 422 mosmol l−1) or in pH (6·78 and 4·79). Further, the surface area of rumen papillae was determined. The feeding of increasing proportions of concentrate resulted in an increase of the surface area of rumen papillae and in an elevated net absorption of sodium, chloride and magnesium. Hypertonicity (422 mosmol l−1) of the artificial rumen fluid caused net influx of water into the rumen but did not influence net absorption of the electrolytes irrespective of the feeding regimen. When the pH was lowered this led to a decrease in net absorption of sodium, chloride and magnesium and in transmural potential difference. The extent of the pH-induced decrease in net ion transport and potential difference was less in sheep receiving high-concentrate diets. When only hay was fed after the period of feeding 90% concentrate diets the surface area of the papillae, the absorptive capacity of the rumen epithelium and the resistance against low pH returned to control levels. The findings show a reversible adaptive response of rumen epithelial functions to different diets.

The effect of diet, intraruminal pH and osmolarity on sodium, chloride and magnesium absorption from the temporarily isolated and washed reticulo-rumen of sheep.

Summary

The effects of lactate, osmotic pressure and/or pH on net absorption of Na, Cl, Mg and water and on transepithelial potential difference were determined in the temporarily isolated reticulo-rumen of sheep. Lactate (0 vs 10 mmol/l) did not change the net absorption of Na, Cl, Mg and water. Hypertonicity (422 mOsm/l) caused a net influx of water but did not influence the net transport of Na, Cl and Mg or the potential difference. A low pH (4.79 vs 6.78) decreased the net absorption of Na by 75% and of Cl by 52% and led to a net secretion of Mg into the rumen. The effects of a low pH were intensified by simultaneous application of lactate and increased osmolarity. It is concluded that ruminal acidosis impairs the absorptive mechanisms of the ruminal epithelium.



Zusammenfassung

Der Einflus von osmotischem Druck, Milchsaure und pH auf die Ionen- und Flussigkeitsabsorption aus dem gewaschenen und zeitweise isolierten Reticulo-Rumen des Schafes



Mit Hilfe des gewaschenen, vorubergehend isolierten Pansens wurden an Schafen die Nettoabsorption von Na, Cl, Mg und Wasser und die transepitheliale Potentialdifferenz bestimmt. In der kunstlichen Pansenflussigkeit wurden Laktatkonzentration, Osmolaritat und pH variiert. Eine Erhohung der Laktatkonzentration von 0 auf 10 mmol/l beeinfluste die Nettoabsorption von Na, Cl, Mg und Wasser nicht. Eine Erhohung der Osmolaritat von 315 auf 422 mOsm/l fuhrte zwar zu einer signifikanten Nettowassersekretion, veranderte aber die Nettoresorption von Na, Cl und Mg und die transepitheliale Potentialdifferenz nicht. Eine pH-Erniedrigung von 6,78 auf 4,79 erniedrigte die NaNettoresorption um 72% und die Cl-Nettoresorption um 52%. Die Mg-Nettoresorption wurde in eine Nettosekretion umgekehrt. Die negativen Effekte auf die Resorption wurden verstarkt, wenn ein niedriger pH-Wert mit erhohter Osmolaritat kombiniert wurde. Die Ergebnisse deuten darauf hin, das wahrend der Pansenacidose die Transportmechanismen der Pansenwand beeintrachtigt sind.

Margret Suendermann

Papers

The effect of diet, intraruminal pH and osmolarity on sodium, chloride and magnesium absorption from the temporarily isolated and washed reticulo-rumen of sheep.

The influence of osmotic pressure, lactic acid and pH on ion and fluid absorption from the washed and temporarily isolated reticulo-rumen of sheep.