Defining comparative physiology: results from an online survey and systematic review.

August Krogh's 1929 principle is referenced as the cornerstone of comparative physiology (CP). However, there are diverse views as to what type of research falls under the CP approach. This study had three aims: 1) determine how CP is defined through an online survey (OS) of physiologists and a systematic review (SR), 2) put forth an updated definition of CP by summarizing OS and SR results, and 3) outline the numerous CP research approaches. Professional physiology societies (n = 54) were invited to share the OS with their members, and a SR was conducted, which yielded 197 and 70 definitions, respectively. The three most common words in descending order in the OS definitions were "different," "animals," and "species" and in the SR definitions, "animals," "species," and "organisms." The three most prevalent themes from the OS and SR definitions were comparing/differences/diversity across species (78% and 51%, respectively), response to the environment/ecology (28% and 43%, respectively), and included evolution or adaptation (24% and 60%, respectively). Ten research approaches were identified, which include broad comparison (i.e., many species generalization), specific comparison (e.g., 2 species; for traits that are different, exaggerated, extreme, missing, or not induced), or comparison while considering evolution (i.e., evolutionary physiology), ecology (i.e., ecophysiology), or human physiology/medicine. Only 5% and 33% of OS and SR definitions described or mentioned Krogh's principle. In conclusion, CP can best be defined as a compilation of research approaches that utilize different types of comparisons to elucidate physiological mechanisms and not simply comparing physiologies as the name implies.

Hibernation induces glutathione redox imbalance in ground squirrel intestine.

Glutathione (GSH) is the major thiol-disulfide redox buffer in cells and is a critical component of antioxidant defense. Here we examined GSH redox balance in the intestinal mucosa during the annual cycle of 13-lined ground squirrels (Spermophilus tridecemlineatus). The ratio of reduced GSH to its oxidized form (glutathione disulfide, GSSG), which is an index of oxidative stress, was five-fold lower in hibernating compared with summer-active squirrels, an effect due primarily to elevated GSSG concentration in hibernators. During hibernation the total pool of GSH equivalents was lowest in squirrels undergoing arousal and highest in squirrels during interbout arousals. Hibernation decreased intestinal GSSG reductase activity by approximately 50%, but had no effect on activities of glutathione peroxidase or glucose-6-phosphate dehydrogenase. Within the hibernation season, expression of the stress protein HSP70 in intestinal mucosa was highest in squirrels entering torpor and early in a torpor bout, and lowest in squirrels arousing from torpor and during interbout euthermia. The results suggest that hibernation in ground squirrels is associated with a shift in intestinal GSH redox balance to a more oxidized state. Higher levels of HSP70 during the early phases of torpor may reflect induction of the stress response due to aberrations in protein folding or may be a mechanism to increase enterocyte tolerance to subsequent stress imposed by extended torpor or the arousal process.

D-glucose releases 5-hydroxytryptamine from human BON cells as a model of enterochromaffin cells.

BACKGROUND & AIMS: 5-Hydroxytryptamine (5-HT) is released from enterochromaffin cells and activates neural reflex programs regulating motility and secretion. Although sugars are reported to release 5-HT in vivo, it is unclear whether they act directly on enterochromaffin cells or indirectly through an intermediary messenger. The aim was to determine if D-glucose is a stimulus for 5-HT release. METHODS: Human BON cells, derived from enterochromaffin cells, were treated with D-glucose, galactose, and the nonmetabolizable methyl alpha-D-glucopyranoside, or with fructose. RESULTS: Reverse-transcription polymerase chain reaction together with Western blot analysis revealed an SGLT-like protein. D-glucose caused a concentration-dependent increase in 5-HT release, which was mimicked by methyl alpha-D-glucopyranoside and galactose but not fructose. D-glucose-stimulated 5-HT release was significantly reduced by phloridzin. Concentrations of mannitol below 75 mmol/L were ineffective in releasing 5-HT. Brefeldin A abolished forskolin-stimulated 5-HT release without affecting basal or constitutive release. CONCLUSIONS: The results show that high concentrations of metabolizable and nonmetabolizable hexoses activate signal transduction pathways, leading to release of 5-HT. These findings imply a role for enterochromaffin cells as "glucose sensors" during ingestion of a meal.

Reactive nitrogen species inhibit alveolar epithelial fluid transport after hemorrhagic shock in rats.

Our recent experimental work demonstrated that a neutrophil-dependent inflammatory response in the lung prevented the normal up-regulation of alveolar fluid clearance by catecholamines following hemorrhagic shock. In this study, we tested the hypothesis that the release of NO within the airspaces of the lung was responsible for the shock-mediated failure of the alveolar epithelium to respond to catecholamines in rats. Hemorrhagic shock was associated with an inducible NO synthase (iNOS)-dependent increase in the lung production of NO and a failure of the alveolar epithelium to up-regulate vectorial fluid transport in response to beta-adrenergic agonists. Inhibition of iNOS restored the normal catecholamine-mediated up-regulation of alveolar liquid clearance. Airspace instillation of dibutyryl cAMP, a stable analog of cAMP, restored the normal fluid transport capacity of the alveolar epithelium after prolonged hemorrhagic shock, whereas direct stimulation of adenyl cyclase by forskolin had no effect. Pretreatment with pyrrolidine dithiocarbamate or sulfasalazine attenuated the iNOS-dependent production of NO in the lung and restored the normal up-regulation of alveolar fluid clearance by catecholamines after prolonged hemorrhagic shock. Based on in vitro studies with an alveolar epithelial cell line, A549 cells, the effect of sulfasalazine appeared to be mediated in part by inhibition of NF-kappaB activation, and the protective effect was mediated by the inhibition of IkappaBalpha protein degradation. In summary, these results provide the first in vivo evidence that NO, released within the airspaces of the lung probably secondary to the NF-kappaB-dependent activation of iNOS, is a major proximal inflammatory mediator that limits the rate of alveolar epithelial transport after prolonged hemorrhagic shock by directly impairing the function of membrane proteins involved in the beta-adrenergic receptor-cAMP signaling pathway in alveolar epithelium.

Involvement of PI 3-kinase in IGF-I stimulation of jejunal Na+-K+-ATPase activity and nutrient absorption.

Mechanisms responsible for increased jejunal transport rates observed in tissues treated with orally administered insulin-like growth factor-I (IGF-I) were studied in 5-day-old colostrum-deprived piglets. Human recombinant IGF-I (3.5 mg. kg(-1). day(-1)) or control vehicle was given orogastrically for 4 days. Disaccharidase activity, fructose uptake, and Na+-glucose cotransporter SGLT-1 protein abundance were similar between groups. Oral IGF-I produced greater rates of enterocyte Na+-K+-ATPase activity with no significant differences in Na+-K+-ATPase abundance. Cellular mechanisms responsible for transport changes were studied in Ussing chambers. In control tissues, the presence of IGF-I in mucosal solutions increased basal short-circuit current (I(sc)), potential difference, D-glucose-stimulated I(sc), and Na+-K+-ATPase activity; these changes were abolished by preincubation of tissues with wortmannin, a phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor. The results suggest that the effect of IGF-I on jejunal ion and nutrient transport involves activation of PI 3-kinase and stimulation of Na+-K+-ATPase activity in enterocytes.

Hibernation induces oxidative stress and activation of NK-kappaB in ground squirrel intestine.

Dramatic changes in blood flow occur during torpor-arousal cycles in mammalian hibernators that could increase the risk of oxidative stress to sensitive tissues. We used 13-lined ground squirrels (Spermophilus tridecemlineatus) to determine the effect of hibernation on lipid peroxidation and expression of stress-activated signaling pathways in the intestine, a tissue highly susceptible to ischemia-reperfusion injury. Compared with summer-active squirrels, levels of the mitochondrial stress protein GRP75 were consistently higher in intestinal mucosa of hibernators in each of five hibernation states (entrance, short-bout torpid, long-bout torpid, arousal and interbout euthermia). The redox-sensitive transcription factor, nuclear factor-kappaB (NF-kappaB), was strongly activated in each hibernation state compared with summer squirrels except for squirrels during an arousal from torpor. In contrast, NF-kappaB activation in brown adipose tissue (BAT) was low in active and hibernating squirrels regardless of season. Levels of conjugated dienes (products of lipid peroxidation) were higher in intestine of hibernators entering torpor and early in a torpor bout compared with summer squirrels. Conjugated diene levels were also higher in short-bout torpid vs arousing squirrels. The results suggest that the intestinal mucosa is vulnerable to oxidative stress during the hibernation season and in response may activate cellular defense pathways that help minimize severe oxidative damage induced by torpor-arousal cycles.

Enterotrophic effect of insulin-like growth factor-I but not growth hormone and localized expression of insulin-like growth factor-I, insulin-like growth factor binding protein-3 and -5 mRNAs in jejunum of parenterally fed rats.

BACKGROUND: Administration of insulin-like growth factor (IGF)-I, but not growth hormone (GH), stimulates mucosal hyperplasia in surgically stressed rats with intestinal atrophy induced by hypocaloric total parenteral nutrition (TPN). Our aim was to characterize the basis for this disparity in enterotrophic action by assessing the relationships between stimulation of intestinal growth, nutritional adequacy, and localization of expression of IGF-I, insulin-like growth factor binding protein (IGFBP)-3 and IGFBP-5 mRNAs in jejunum. METHODS: Rats were maintained with TPN for 8 days and treated with IGF-I or GH and adequate nutrition for 5 days after recovery from surgery. Jejunal mass, morphology, and sucrase activity were assessed. Localization of expression of IGF-I, IGFBP-3, and IGFBP-5 mRNAs in jejunum was accomplished by in situ hybridization. RESULTS: Serum IGF-I and body weight gain were significantly increased by IGF-I or GH. Jejunal mucosal dry mass, morphology, and sucrase activity were improved with IGF-I but not GH. There were no differences in IGF-I mRNA. IGFBP-3 mRNA was localized in the lamina propria of the villi. IGF-I or GH stimulated IGFBP-3 expression. IGF-I strongly stimulated IGFBP-5 expression in the lamina propria and the muscularis and induced a twofold increase in IGFBP-5 mRNA based on RNase protection assay of intact jejunum total RNA. GH induced a modest increase in IGFBP-5 expression in the muscularis with no effect on intact jejunum total RNA. CONCLUSIONS: The GH resistance observed in the jejunal mucosa of TPN rats cannot be fully explained by inadequate nutrition. The expression of IGFBP-5 in the lamina propria suggests it may modulate the enterotrophic action of exogeneous IGF-I.

Intestinal transport during fasting and malnutrition.

Fasting or malnutrition (FM) has dramatic effects on small intestinal mucosal structure and transport function. Intestinal secretion of ions and fluid is increased by FM both under basal conditions and in response to secretory agonists. Intestinal permeability to ions and macromolecules may also be elevated by FM, which increases the potential for fluid and electrolyte losses and for anaphylactic responses to luminal antigens. Mucosal atrophy induced by FM reduces total intestinal absorption of nutrients, but nutrient absorption normalized to mucosal mass may actually be enhanced by a variety of mechanisms, including increased transporter gene expression, electrochemical gradients, and ratio of mature to immature cells. These observations underscore the value of enteral feeding during health and disease.

Neural control of intestinal ion transport and paracellular permeability is altered by nutritional status.

This study examined the effect of fasting on the neural control of ion transport and paracellular permeability in piglet jejunum. Muscle-stripped tissues from fed or 48-h fasted piglets were mounted in Ussing chambers. Neural blockade with tetrodotoxin (TTX) or antagonists of muscarinic or nicotinic receptors caused reductions in basal short-circuit current that were approximately threefold greater in fasted piglets. The TTX-induced reduction in short-circuit current in fasted piglets was due to a decrease in residual ion flux and was abolished in the absence of HCO(-)(3). Intestinal paracellular permeability, as indicated by tissue conductance (G(t)) and fluxes of inulin and mannitol, was significantly increased by fasting. TTX increased inulin flux and G(t) in fed but not fasted piglets. In fasted piglets, carbachol reduced G(t) by 29% and mannitol flux by 27% but had no effect on these parameters in the fed state. We conclude that fasting enhances enteric neural control of basal ion transport and increases paracellular permeability in piglet jejunum. Tonic release of enteric neurotransmitters regulates paracellular permeability in the fed state, and cholinergic stimulation restores fasting-induced elevations in paracellular permeability to fed levels.

Differential jejunal and colonic adaptation due to resection and IGF-I in parenterally fed rats.

Patients with severe short-bowel syndrome (SBS) often require long-term total parenteral nutrition (TPN) to maintain their nutritional status because of limited intestinal adaptation. Growth factors, including insulin-like growth factor I (IGF-I), are under investigation to promote intestinal adaptation and tolerance to oral feeding. We investigated structural and functional adaptation of the jejunum and colon in four groups of rats maintained with TPN for 7 days after a 60% jejunoileal resection and cecectomy or sham surgery and treatment with IGF-I or vehicle. Resection alone did not stimulate jejunal growth. IGF-I significantly increased jejunal mucosal mass, enterocyte proliferation, and migration rates. IGF-I decreased jejunal sucrase specific activity and reduced active ion transport and ionic permeability; resection alone had no effect. In contrast, resection significantly increased colonic mass and crypt depth but had no effect on active ion transport or ionic permeability. IGF-I had minimal effects on colonic structure. IGF-I but not resection stimulates jejunal adaptation, whereas resection but not IGF-I stimulates colonic growth in rats subjected to a model for human SBS. IGF-I treatment may improve intestinal adaptation in humans with SBS.

Oral IGF-I enhances nutrient and electrolyte absorption in neonatal piglet intestine.

The effect of orally administered insulin-like growth factor-I (IGF-I) on small intestinal structure and function was studied in 5-day-old colostrum-deprived piglets. Human recombinant IGF-I (3.5 mg. kg(-1). day(-1)) or control vehicle was given orogastrically for 4 days. Body weights, jejunal and ileal mucosa wet and dry weights, and serum IGF-I levels were similar in the two groups. Small intestinal villus height and crypt depth and jejunal enterocyte microvillar dimensions were also similar between groups. Oral IGF-I produced higher rates of jejunal ion transport because of increased basal Na+ absorption. Short-circuit current responses to mucosal addition of D-glucose and L-alanine and net transepithelial absorption of 3-O-methylglucose were increased by IGF-I. Carrier-mediated uptake of D-glucose per milligram in everted jejunal sleeves was greater in IGF-I-treated piglets because of a significantly greater maximal rate of uptake. We conclude that rates of net Na+ and Na+-dependent nutrient absorption are enhanced in piglets treated with oral IGF-I, and this effect is independent of changes in mucosal mass or surface area.

Psyllium improves fecal consistency and prevents enhanced secretory responses in jejunal tissues of piglets infected with ETEC.

Infection with enterotoxigenic E. coli (ETEC) induces secretory diarrhea by stimulating net secretion of fluid and electrolytes. We tested the hypothesis that ETEC potentiates jejunal ion secretion induced by other agonists and also examined whether the soluble fiber psyllium ameliorates effects of ETEC-induced pathophysiology. Noninfected or ETEC-infected piglets were given oral electrolyte solution twice daily or electrolyte solution supplemented with psyllium for 48 hr. Jejunal tissues were mounted in flux chambers and basal and stimulated ion transport responses, as reflected by short-circuit current (I(SC)) were measured. The severity of ETEC-induced diarrhea was reduced by psyllium. I(SC) responses to carbachol and 5-hydroxytryptamine were greater in tissues from infected piglets compared with noninfected controls or infected piglets given psyllium. These results suggest that psyllium ameliorates ETEC-induced diarrhea and prevents the enhanced secretory responses to calcium-mediated agonists that occur in ETEC-infected piglet jejunum.

Reduced susceptibility of mice overexpressing transforming growth factor alpha to dextran sodium sulphate induced colitis.

BACKGROUND: Transforming growth factor alpha (TGF-alpha) knockout mice have increased susceptibility to dextran sodium sulphate (DSS) induced colitis. AIM: To substantiate the findings that TGF-alpha is a key mediator of colonic mucosal protection and/or repair mechanisms by evaluating the susceptibility of mice overexpressing TGF-alpha to DSS induced colitis. METHODS: TGF-alpha overexpression was induced in transgenic mice by ZnSO4 administration in drinking water (TG+). Three groups were used as controls: one transgenic group without ZnSO4 administration (TG-), and two non-transgenic littermate groups receiving ZnSO4 (Non-TG+) or only water (Non-TG-). Acute colitis was induced in all groups by administration of DSS (5%, w/v) in drinking water for six days and libitum. RESULTS: About 35-39% of the entire colonic mucosa was destroyed in Non-TG-, Non-TG+, and TG- animals compared with 9% in TG+ mice. the crypt damage score was 18.7 (0.9), 18.2 (1.0), 18.9 (0.8), and 6.8 (1.5) (means (SEM)) in Non-TG-, Non-TG+, TG-, and TG+ mice respectively. Mucin and bromodeoxyuridine staining were markedly enhanced in colons of TG+ mice compared with controls, indicating increased mucosal protection and regeneration. CONCLUSIONS: The significantly reduced susceptibility of mice overexpressing TGF-alpha to DSS further substantiates that endogenous TGF-alpha is a pivotal mediator of protection and/or healing mechanisms in the colon.

Hibernation induces expression of moesin in intestinal epithelial cells.

Identification of proteins that are differentially expressed in mammals that hibernate can provide insight into mechanisms that preserve cellular function at low temperatures. A candidate protein was identified in intestinal brush border membranes of 13-lined ground squirrels. Intestinal brush border membrane proteins were separated using SDS-PAGE and gels were stained with Coomassie blue. We observed a approximately 75-kDa band that was specifically increased in brush border membranes isolated from torpid squirrels compared with summer active squirrels. The 75-kDa band was cut from one-dimensional gels and sequenced. A 17 amino acid sequence was identified of which amino acids 2-17 matched exactly a portion of moesin, a membrane-cytoskeletal linking protein and member of the ERM (ezrin/radixin/moesin) family. The sequence results were confirmed using anti-moesin antibodies that detected strong bands at approximately 75 kDa on Western blots of brush border membranes in torpid squirrels (Tb approximately 7 degreesC) and only faint signals in summer squirrels (Tb approximately 37 degrees C) or aroused hibernators (Tb approximately 37 degrees C). In contrast, signals obtained using anti-ezrin antibodies were uniformly strong in all squirrels, regardless of activity state. Intestinal brush borders of mice and rats expressed ezrin but not moesin. These results provide evidence for the physiological induction of an ERM protein in intestinal epithelial cells of torpid hibernators and support the idea that hibernation involves differential expression of gene products that may facilitate viability of cells at low temperatures.

Muscarinic inhibition of substance P induced ion secretion in piglet jejunum.

We examined the effects of the muscarinic agonist carbachol on ion secretion induced by substance P (SP) in piglet jejunal tissues mounted in Ussing chambers. Tetrodotoxin was present in all solutions to inhibit neural activity. Carbachol added 10 min prior to 0.75 microM SP dose dependently inhibited subsequent SP responses, with 90% inhibition at 10 microM carbachol. Addition of an equipotent dose of SP (7.5 microM) had no effect on subsequent carbachol-induced secretion. Carbachol's inhibition of SP-induced secretion was evident for at least 45 min and was abolished by prior addition of the M3 receptor antagonist 4-diphenylacetoxy-N-methyl-piperidine methiodide (4-DAMP), but remained intact in the presence of the M2 antagonist gallamine or the nicotinic antagonist mecamylamine. Atropine added 10 min after carbachol restored subsequent SP responses toward control levels. Carbachol also reduced secretory responses to histamine and, to a lesser extent, prostaglandin E2 (PGE2). SP-induced secretion was not affected by prior addition of histamine and was reduced by PGE2 only at the highest PGE2 concentration. The results suggest that activation of the epithelial M3 receptor by carbachol inhibits subsequent secretory responses to the calcium-mediated agonists SP and histamine in piglet jejunum. This may reflect muscarinic activation of a negative messenger in epithelial cells that limits Cl- secretion.

Stimulation of intestinal growth is associated with increased insulin-like growth factor-binding protein 5 mRNA in the jejunal mucosa of insulin-like growth factor-I-treated parenterally fed rats.

Surgically stressed rats maintained with total parenteral nutrition (TPN) exhibit jejunal atrophy, which can be attenuated by insulin-like growth factor-I (IGF-I) but not by growth hormone (GH) treatment. In order to understand the basis for the selective action of IGF-I, the levels of mRNAs encoding IGF-I, IGF-binding proteins (IGFBPs), IGF-I receptor, and GH receptor/binding protein (GHR/GHBP) were determined in rats given TPN and treated with GH, IGF-I, or GH + IGF-I. GH treatment significantly stimulated hepatic IGF-I mRNA. IGF-I treatment did not alter liver IGF-I mRNA, nor was there any evidence for interaction between GH and IGF-I. Jejunal mucosa IGF-I mRNA was extremely low and was not altered by TPN or by any of the hormonal treatments. The inability of GH to stimulate jejunal growth was not associated with a deficiency in GHR/GHBP mRNA. In jejunal mucosa, IGF-I and GH treatment independently and synergistically stimulated IGFBP-3 mRNA. IGF-I stimulated jejunal IGFBP-5 mRNA, but GH had no effect on IGFBP-5 mRNA. The levels of IGF-I receptor and IGFBP-1, 2, 4, and 6 mRNAs were extremely low and/or were not altered by any of the treatments. These results suggest that the ability of exogenous IGF-I, but not GH, to induce IGFBP-5 mRNA in jejunal mucosa may lead to the selective growth-promoting effect of IGF-I. Jejunal mucosa IGFBP-3 mRNA levels were not correlated with altered growth. We postulate that IGFBP-5 positively modulates the anabolic effects induced by exogenous IGF-I in the jejunum.