Ca-dependent slow action potentials in human skeletal muscle

Slow Ca-action potentials (CaAP) were studied in normal human skeletal muscle fibers obtained during surgery (fibers with both ends cut). Control studies also were carried out with intact as well as cut rat skeletal muscle fibers. Experiments were performed in hypertonic Cl-free saline with 10 or 84 mM Ca and K-channel blockers; muscles were preincubated in a saline containing Cs and tetraethylammonium. A current-clamp technique with two intracellular microelectrodes was used. In human muscle, 14.5% of the fibers showed fully developed CaAPs, 21% displayed nonregenerative Ca responses, and 64.5% showed only passive responses; CaAPs were never observed in 10 mM Ca. In rat muscle, nearly 90% of the fibers showed CaAPs, which were not affected by the cut-end condition. Human and rat muscle fibers had similar membrane potential and conductance in the resting state. In human muscle (22-32 degrees C, 84 mM Ca), the threshold and peak potential during a CaAP were +26 +/- 6 mV and +70 +/- 3 mV, respectively, and the duration measured at threshold level was 1.7 +/- 0.5 sec. In rat muscle, the duration was four times longer. During a CaAP, membrane conductance was assumed to be a leak conductance in parallel with a Ca and a K conductance. In human muscle (22-32 degrees C, 84 mM Ca, 40 micron fiber diameter), values were 0.4 +/- 0.1 microS, 1.1 +/- 0.7 microS, and 0.9 +/- 0.4 microS, respectively. Rat muscle (22-24 degrees C, 84 mM Ca) showed leak and K conductances similar to those found in human fibers. Ca-conductance in rat muscle was double the values obtained in human muscle fibers.     

The K(+) affinity of gastric H(+),K(+)-ATPase is affected by both lipid composition and the beta-subunit

It is generally assumed that negatively charged residues present in the alpha-subunit of gastric H(+),K(+)-ATPase are involved in K(+) binding and transport. Despite the fact that there is no difference between various species regarding these negatively charged residues, it was observed that the apparent K(+) affinity of the pig enzyme was much lower than that of the rat H(+),K(+)-ATPase. By determining the K(+)-stimulated dephosphorylation reaction of the phosphorylated intermediate K(0.5) values for K(+) of 0.12+/-0.01 and 1.73+/-0.03 mM were obtained (ratio 14.4) for the rat and the pig enzyme, respectively. To investigate the reason for the observed difference in K(+) sensitivity, both enzymes originating from the gastric mucosa were either reconstituted in a similar lipid environment or expressed in Sf9 cells. After reconstitution in K(+)-permeable phosphatidylcholine/cholesterol liposomes K(0.5) values for K(+) of 0.16+/-0.01 and 0.35+/-0.05 mM for the rat and pig enzyme respectively were measured (ratio 2.2). After expression in Sf9 cells the pig gastric H(+),K(+)-ATPase still showed a 4.1 times lower K(+) sensitivity than that of the rat enzyme. This means that the difference in K(+) sensitivity of the rat and pig gastric H(+), K(+)-ATPase is not only due to a different lipid composition but also to the structure of either the alpha- or beta-subunit. Expression of hybrid enzymes in Sf9 cells showed that the difference in K(+) sensitivity between the rat and pig gastric H(+),K(+)-ATPase is primarily due to differences in the beta-subunit.     

Identification of a VIP-specific receptor in guinea pig tenia coli

Vasoactive intestinal polypeptide (VIP) and pituitary adenylate cyclase-activating peptide (PACAP) interact with VPAC(2) receptors in rabbit and guinea pig (GP) gastric muscle but with functionally distinct VIP and PACAP receptors in GP tenia coli. This study examined whether selectivity for VIP was determined by two residues (40, 41) in the extracellular domain that differ in the VIP receptors of GP gastric and tenial muscle. A mutant rat VPAC(2) receptor (L40F, L41F), and two chimeric receptors in which the NH(2)-terminal domain of rat VPAC(2) receptor was replaced with that of GP gastric (chimeric-G) or tenia coli (chimeric-T) VIP receptors, were constructed and expressed in COS-1 cells. VIP and PACAP bound with equal affinity to wild-type and mutant rat VPAC(2) receptors and to chimeric-G receptor (IC(50): VIP 0.3 +/- 0.1 to 1.5 +/- 0.4 nM, PACAP 0.4 +/- 0.1 to 2.5 +/- 0.1 nM) and stimulated cAMP with equal potency (EC(50): VIP 13 +/- 5 to 48 +/- 8 nM, PACAP 8 +/- 3 to 31 +/- 14 nM). VIP bound with high affinity also to chimeric-T receptor (IC(50): 0.5 +/- 0.1 nM) and stimulated cAMP with high potency (EC(50): 3 +/- 1 nM). In contrast, PACAP exhibited >1,000-fold less affinity for binding or potency for stimulating cAMP. We conclude that GP tenia coli express a VIP-specific receptor and that selectivity is determined by a pair of extracellular phenylalanine residues.     

Ammonia-utilizing enzymes of adherent bacteria in the sheep's rumen

In experiments on 6 sheep the authors found the following enzyme activities in bacteria in the rumen fluid, bacteria adhering to the epithelium of the rumen wall and bacteria adhering to food particles in the rumen (given in nkat X g-1 bacterial dry weight): GDH (NADH): 725 +/- 165, 558 +/- 127, 661 +/- 153; GDH (NADPH): 558 +/- 338, 255 +/- 88, 565 +/- 139; GOAT (NADH): 46 +/- 23, 67 +/- 31, 66 +/- 14; GOGAT/NADPH: 58 +/- 27, 56 +/- 15, 65 +/- 29; GS: 153 +/- 65, 69 +/- 35, 71 +/- 32; ALT: 71 +/- 25, 43 +/- 20, 52 +/- 11; AST: 52 +/- 12, 33 +/- 16, 28 +/- 15. The results show that, except for GDH (NADPH), there were no significant differences between the given enzyme activities in the rumen fluid and in bacteria adhering to the rumen wall and to food. Adherent rumen bacteria have the same potential possibilities as the rumen fluid bacteria for the utilization of ammonia, particularly for the synthesis of glutamic acid, glutamine, alanine and aspartic acid, with the above enzymes as catalysts. By means of the GS/GOGAT system, adherent rumen bacteria can probably synthesize glutamic acid in the presence of a limited NH3 concentration in the rumen.     

Characterisation of gastric ghrelin cells in man and other mammals: studies in adult and fetal tissues

Ghrelin is a new gastric peptide involved in food intake control and growth hormone release. We aimed to assess its cell localisation in man during adult and fetal life and to clarify present interspecies inconsistencies of gastric endocrine cell types. A specific serum generated against amino acids 13-28 of ghrelin was tested on fetal and adult gastric mucosa and compared with ghrelin in situ hybridisation. Immunogold electron microscopy was performed on normal human, rat and dog adult stomach. Ghrelin cells were detected in developing gut, pancreas and lung from gestational week 10 and in adult human, rat and dog gastric mucosa. By immunogold electron microscopy, gastric ghrelin cells showed distinctive morphology and hormone reactivity in respect to histamine enterochromaffin-like, somatostatin D, glucagon A or serotonin enterochromaffin cells. Ghrelin cells were characterised by round, compact, electron-dense secretory granules of P/D(1) type in man (mean diameter 147+/-30 nm), A-like type in the rat (183+/-37 nm) and X type in the dog (273+/-49 nm). It is concluded that, ghrelin is produced by well-defined cell types, which in the past had been labelled differently in various mammals mostly because of the different size of their secretory granule. In man ghrelin cells develop during early fetal life.     

In vitro metabolism and permeation studies in rat jejunum: organic chromium compared to inorganic chromium.

The purpose of these studies was to compare the in vitro absorption of two inorganic chromium(III) compounds: chromium chloride and chromium nitrate, with organic chromium(III)-picolinate; and to investigate if any in vitro metabolism of chromium(VI) takes place. The in vitro metabolism studies showed that chromium (VI) was reduced by artificial gastric juice. The reduction followed first order kinetics with a half-life of 23 min. The studies also showed that the chromium picolinate complex was stable in artificial gastric juice for 4 hours. By the rat everted gut sac technique, chromium chloride, chromium nitrate and chromium picolinate penetrated the rat jejunum with 165 +/- 59, 160 +/- 26 and 127 +/- 36 ng chromium per g rat jejunum, respectively, whereas the permeability coefficients (Papp) were 0.7 +/- 0.3, 1.0 +/- 0.4, and 9.6 +/- 2.2 microns/min, respectively. Absorption studies on pig intestine in Ussing chambers showed a nearly total adsorption of chromium(III) by the chambers, resulting in unreliable data.     

Characterization of pancreas-projecting rat dorsal motor nucleus of vagus neurons

The electrophysiological and morphological properties of rat dorsal motor nucleus of the vagus (DMV) neurons innervating the pancreas were examined by using whole cell patch clamp recordings from brain stem slices and postfixation morphological reconstructions of Neurobiotin-filled neurons. Recordings were made from 178 DMV neurons whose projections had been identified by previous apposition of the fluorescent neuronal tracer DiI to the body of the pancreas. DMV neurons projecting to the pancreas had an input resistance of 434 +/- 14 M omega, an action potential duration of 3 +/- 0.1 ms, and an afterhyperpolarization of 18 +/- 0.4 mV amplitude and 108 +/- 7 ms time constant of decay; these electrophysiological properties resembled those of gastric-projecting neurons but were significantly different from those of intestinal-projecting neurons. Interestingly, 14 of 178 pancreas-projecting neurons showed the presence of a slowly developing afterhyperpolarization whose presence was not reported in DMV neurons projecting to any other gastrointestinal area. The morphological characteristics of pancreas-projecting neurons (soma area 274 +/- 12 microm2; soma diameter of 25 +/- 0.7 microm; soma form factor 0.74 +/- 0.01; segments 9.7 +/- 0.41), however, were similar to those of intestinal- but differed from those of gastric-projecting neurons. In summary, these results suggest that pancreas-projecting rat DMV neurons are heterogeneous with respect to some electrophysiological and morphological properties. These differences might underlie functional differences in the vagal modulation of pancreatic functions.     

Caco-2 cells in culture synthesize and degrade dopamine and 5-hydroxytryptamine: a comparison with rat jejunal epithelial cells

To explore the usefulness of Caco-2 cells in the study of intestinal dopaminergic and 5-hydroxytryptaminergic physiology, we have undertaken the study of aromatic L-amino acid decarboxylase (AADC), catechol-O-methyltransferase (COMT) and type A and B monoamine oxidase (MAO-A and MAO-B) activities in these cells using specific substrates. The activity of these enzymes was also evaluated in isolated rat jejunal epithelial cells. The results showed that Vmax values (in nmol mg protein(-1) h(-1)) for AADC, using L-DOPA as the substrate, in rat jejunal epithelial cells (127.3+/-11.4) were found to be 6-fold higher than in Caco-2 cells (22.5+/-2.6). However, Km values in Caco-2 cells (1.24+/-0.37 mM) were similar to those observed in rat jejunal epithelial cells (1.30+/-0.29 mM). Similar results were obtained when AADC activity was evaluated using L-5HTP as substrate; in rat jejunal epithelial cells Vmax values (in nmol mg prot(-1) h(-1)) were found to be 5-fold that in Caco-2 cells (16.3+/-1.0 and 3.0+/-0.2, respectively), and Km values in Caco-2 cells (0.23+/-0.08 mM) were again similar to those observed in rat intestinal epithelial cells (0.09+/-0.03 mM). Caco-2 cells were not able to O-methylate dopamine, in contrast to rat jejunal epithelial cells (Vmax = 8.6+/-0.4 nmol mg protein(-1)(h-1); Km = 516+/-57 microM). Vmax values (in nmol mg protein(-1)(h-1)) for type A and B MAO in Caco-2 cells (19.0+/-0.6 and 5.4+/-0.6, respectively) were found to be significantly lower (P<0.05) than those in rat jejunal epithelial cells (46.9+/-3.1 and 9.6+/-1.2, respectively); however, no significant differences in the Km values were observed between Caco-2 and rat jejunal epithelial cells for both type A and B MAO. In conclusion, Caco-2 cells in culture are endowed with the synthetic and metabolic machinery needed to form and degrade DA and 5-HT, though, no COMT activity could be detected in these cells.     

In vivo microdialysis for determination of nasal liquid ion composition

Airway surface liquid (ASL) contains substances important in mucociliary clearance and airway defense. Little is known about substance concentrations in ASL because of its small volume and sampling difficulties. We used in vivo microdialysis (IVMD) to sample liquid lining the nasal cavity without net volume removal and incorporated into IVMD a potential difference (PD) electrode to assess airway integrity. The cystic fibrosis (CF) mouse nasal epithelia exhibit ion transport defects identical to those in CF human airways and, thus, are a good model for CF disease. We determined that nasal liquid [Na+] (107 +/- 4 mM normal; 111 +/- 9 mM CF) and [Cl-] (120 +/- 6 mM normal; 122 +/- 4 mM CF) did not differ between genotypes. The nasal liquid [K+] (8.7 +/- 0.4 mM) was significantly less in normal than in CF mice (16.6 +/- 4 mM). IVMD accurately samples nasal liquid for ionic composition. The ionic composition of nasal liquid in the normal and CF mice is similar.     

Blood acid-base status, whole blood and whole body buffer values in sand rats (Psammomys obesus) exposed to hypercapnia

Arterial blood acid-base status of unanesthetized sand rats (Psammomys obesus) were studied under normocapnic and hypercapnic conditions, and compared to those obtained for the albino rat (Rattus norvegicus). The average control blood pH: 7.396 +/- 0.034; PaCO2: 30.5 +/- 2.9 mmHg; HCO-3: 18.8 +/- 2.5 mM/l; and HCO-3 std: 20.9 +/- 2.1 (N = 15) obtained here for the sand rat are in the lower range of values found in other mammals and indicate a status of partially compensated metabolic acidosis. The blood buffer values of the sand rat, delta log PCO2/delta pH = -2.32 +/- 0.35 (N = 25) are significantly higher than those found here for the rat, delta log PCO2/delta pH = -1.51 +/- 0.10 (N = 39), and those reported for other mammals. This high blood buffer value may be related to the natural high mineral diet of the sand rat. The in vivo (whole body) buffer value delta log PaCO2/delta pH = -1.41 and -1.65 for the sand rat and the rat found here are higher than those reported for the man and dog and may represent a physiological adaptation to the hypercapnic conditions prevailing in underground burrows.     

Differences in intragastric pH in diabetic vs. idiopathic gastroparesis: relation to degree of gastric retention

Evidence suggests that distinct mechanisms underlie diabetic and idiopathic gastroparesis. Differences in gastric acid in gastroparesis of different etiologies and varying degrees of gastric stasis are uninvestigated. We tested the hypotheses that 1) gastric pH profiles show differential alteration in diabetic vs. idiopathic gastroparesis and 2) abnormal pH profiles relate to the severity of gastric stasis. Sixty-four healthy control subjects and 44 gastroparesis patients (20 diabetic, 24 idiopathic) swallowed wireless transmitting capsules and then consumed (99m)Tc-sulfur colloid-labeled meals for gastric scintigraphy. Gastric pH from the capsule was recorded every 5 s. Basal pH was higher in diabetic (3.64 +/- 0.41) vs. control subjects (1.90 +/- 0.18) and idiopathic subjects (2.41 +/- 0.42; P < 0.05). Meals evoked initial pH increases that were greater in diabetic (4.98 +/- 0.32) than idiopathic patients (3.89 +/- 0.39; P = 0.03) but not control subjects (4.48 +/- 0.14). pH nadirs prior to gastric capsule evacuation were higher in diabetic patients (1.50 +/- 0.23) than control subjects (0.58 +/- 0.11; P = 0.003). Four-hour gastric retention was similar in diabetic (18.3 +/- 0.5%) and idiopathic (19.4 +/- 0.5%) patients but higher than control subjects (2.2 +/- 0.5%; P < 0.001). Compared with control subjects, those with moderate-severe stasis (>20% retention at 4 h) had higher basal (3.91 +/- 0.55) and nadir pH (2.23 +/- 0.42) values (P < 0.05). In subgroup analyses, both diabetic and idiopathic patients with moderate-severe gastroparesis exhibited increased pH parameters vs. those with mild gastroparesis. In conclusion, diabetic patients with gastroparesis exhibit reduced gastric acid, an effect more pronounced in those with severely delayed gastric emptying. Idiopathic gastroparetic subjects exhibit nearly normal acid profiles, although those with severely delayed emptying show reduced acid vs. those with mild delays. Thus both etiology and degree of gastric stasis determine gastric acidity in gastroparesis.     

Purification of a nickel-containing urease from the rumen anaerobe Selenomonas ruminantium

Urease was purified 592-fold to homogeneity from the anaerobic rumen bacterium Selenomonas ruminantium. The urease isolation procedure included a heat step and ion-exchange, hydrophobic, gel filtration, and fast protein liquid chromatography. The purified enzyme exhibited a Km for urea of 2.2 +/- 0.5 mM and a Vmax of 1100 mumol of urea min-1 mg-1. The molecular mass estimated for the native enzyme was 360,000 +/- 50,000 daltons, whereas a subunit value of 70,000 +/- 2,000 daltons was determined. These results are in contrast to the findings of Mahadevan et al. (Mahadevan, S., Sauer, F. D., and Erfle, J. D. (1977) Biochem. J. 163, 495-501) in which isolated rumen urease was reported to be one-third this size (Mr 120,000-130,000) and to catalyze urea hydrolysis at a maximum velocity of only 53 mumol min-1 mg-1. S. ruminantium urease contained 2.1 +/- 0.4 nickel ions/subunit, comparable to the nickel content in jack bean urease (Dixon, N.E., Gazzola, C., Blakeley, R.L., and Zerner, B. (1975) J. Am. Chem. Soc. 97, 4131-4133). Thus, the active site of bacterial urease is very similar to that found in the plant enzymes.     

Growth hormone deficiency: permanence and diagnosis in young adults

OBJECTIVE: To optimize the tools for diagnosing idiopathic growth hormone (GH) deficiency. METHODS: We compared the data of 43 young adults treated for GH deficiency before and after GH treatment and puberty. Those with organic lesions were assigned to group 1 (n = 9), those with certain GH deficiency (n = 11) to group 2 and those with no criterion of certitude of GH deficiency to group 3 (n = 23). RESULTS: Group 1 patients: the GH peaks at first [1.5 +/- (SE) 0.4 microg/l] and second (1.9 +/- 0.7 microg/l) evaluations before treatment were similar to those at the third evaluation (1.2 +/- 0.8 microg/l) after treatment. Group 2 patients: they had similar peaks (2.6 +/- 0.8, 2.9 +/- 0.5 and 5.5 +/- 1.4 microg/l). Group 3 patients: the peaks increased from 4.9 +/- 0.4 and 4.8 +/- 0.4 to 18.4 +/- 2.3 microg/l (p < 0.0001); 87% had a GH peak >10 microg/l at this evaluation. The plasma insulin-like growth factor 1 was initially below -2 z-score in 12/13 of these patients and similarly low in 4/17 patients at the third evaluation. The growth rates of the three groups before and their increase during the 1st year of treatment were similar. CONCLUSION: Almost all patients with GH deficiency before puberty without criteria of certitude had a normal GH peak after puberty. Some of these patients probably had a transiently low GH secretion.     

Immunoreactive calmodulin in supernatant fraction of platelet homogenates from humans, guinea pigs, rats, and rabbits

Immunoreactive Calmodulin (CM) was measured in the supernatant fraction of homogenates of platelets obtained from humans, rats, guinea pigs and rabbits, using a commercial RIA kit. The levels (microgram/g wet weight of platelets) of immunoreactive CM were 6.8 +/- 0.5, 6.9 +/- 0.4, 8.6 +/- 1.8 and 9.7 +/- 3.1 (mean +/- SEM) for rat, human, rabbit and guinea pig, respectively. There was no statistically significant difference in values between these four species. The refractoriness of rat platelets to aggregate to certain agonists such as platelet activating factor (PAF) cannot be explained on differences in amount of immunoreactive CM.     

Reduced tLESR elicitation in response to gastric distension in fundoplication patients

Transient lower esophageal sphincter relaxations (tLESRs) are vagally mediated in response to gastric cardiac distension. Nine volunteers, eight gastroesophageal reflux disease (GERD) patients, and eight fundoplication patients were studied. Manometry with an assembly that included a barostat bag was done for 1 h with and 1 h without barostat distension to 8 mmHg. Recordings were scored for tLESRs and barostat bag volume. Fundoplication patients had fewer tLESRs (0.4 +/- 0.3/h) than either normal subjects (2.4 +/- 0.5/h) or GERD patients (2.0 +/- 0.3/h). The tLESRs rate increased significantly in normal subjects (5.8 +/- 0.9/h) and GERD patients (5.4 +/- 0.8/h) during distension but not in the fundoplication group. All groups exhibited similar gastric accommodation (change in volume/change in pressure) in response to distension. Fundoplication patients exhibit a lower tLESR rate at rest and a marked attenuation of the response to gastric distension compared with either controls or GERD patients. Gastric accommodation was not impaired with fundoplication. This suggests that the receptive field for triggering tLESRs is contained within a wider field for elicitation of gastric receptive relaxation and that only the first is affected by fundoplication.     

Marked increase in gastric histidine decarboxylase activity in patients with hypergastrinemia.

Histidine decarboxylase (HDC) activity and histamine content were measured in endoscopic gastric biopsy specimens of 19 control subjects with normogastrinemia and 6 patients with hypergastrinemia. In controls, the HDC activity was 3 fold higher in fundic mucosa (120 +/- 13 fmol/min/mg protein, mean +/- S.E.) than in antral mucosa (39 +/- 5 fmol/min/mg protein). In patients with hypergastrinemia, an extremely high HDC activity (713 +/- 181 fmol/min/mg protein) was observed in fundic mucosa, although the HDC activity in antral mucosa was not significantly different from that of controls. The histamine content in fundic mucosa was also significantly higher in patients with hypergastrinemia than in controls but no significant difference was seen in histamine content in antral mucosa between the two groups. These results are compatible with the hypothesis that in man, as well as in rat, histamine synthesis in fundic mucosa is enhanced by gastrin.     

Identification of receptors in a system of functionally heterogeneous proteins specifically bound to androgens in rat liver cytosol

The methods of androgen receptor (RA) isolation and identification in rat liver cytosol were studied. It was shown that male rat liver contains a system of specific androgen (A)-binding proteins consisting of at least three main components: RA, delta 4-androstendione (delta 4-A)-binding component and an unusual estrogen-binding protein interacting also with A and the first two components in females. The identity of one of A-binding components to RA was proved by cumulative properties of this component which are similar to those of RA from other tissues. These properties are as follows: 1) high values of apparent association constant, Ka, for 3H-R1881 (2.8 +/- 0.3 X 10(8) M-1) and 3H-5 alpha-dihydrotestosterone (3H-DHT) (5.0 +/- 0.4 X 10(8) M-1); 2) low binding capacity--approximately 10 fmol/mg of protein of nonfractionated cytosol; 3) pronounced specificity of affinity for active A (DHT, R1881, testosterone); 4) large size of the protein molecule (6.5 +/- 0.25 nm); 5) ability to decrease this size to 3.2 +/- 0.08 nm in a high ionic strength buffer; 6) precipitation at low concentrations of ammonium sulfate: 7) strong interaction with heparin-Sepharose. The properties of the delta 4-A-binding component do not coincide with those of RA: it has a low Ka for 3H-delta 4-A (1.15 +/- 0.5 X 10(6) M-1), a high binding capacity (1.22 +/- 0,12 pmol/mg of protein of nonfractionated cytosol) and can bind various delta 4-3-ketosteroids irrespective of the degree and nature of their biological activity. It was concluded that preliminary isolation of rat liver RA on heparin-Sepharose can be used for differential identification and characterization of this protein.     

The effect of extracellular potassium on the intracellular potassium ion activity and transmembrane potentials of beating canine cardiac Purkinje fibers

We used open tip microelectrodes containing a K+-sensitive liquid ion exchanger to determine directly the intracellular K+ activity in beating canine cardiac Purkinje fibers. For preparations superfused with Tyrode's solution in which the K+ concentration was 4.0 mM, intracellular K+ activity (ak) was 130.0+/-2.3 mM (mean+/-SE) at 37 degrees C. The calculated K+ equilibrium potential (EK) was -100.6+/- 0.5 mV. Maximum diastolic potential (ED) and resting transmembrane potential (EM) were measured with conventional microelectrodes filled with 3 M KCl and were -90.6+/-0.3 and -84.4+/-0.4 mV, respectively. When [K+]o was decreased to 2.0 mM or increased to 6.0, 10.0, and 16.0 mM, ak remained the same. At [K+]o=2.0, ED was -97.3+/-0.4 and Em - 86.0+/-0.7 mV; at [K+]o=16.0, ED fell to -53.8+/-0.4 mV and Em to the same value. Over this range of values for [K+]o, EK changed from - 119.0+/-0.3 to -63.6+/-0.2 mV. These values for EK are consistent with those previously estimated indirectly by other techniques.     

Skeletal muscle apoptosis is not increased in gastric cancer patients with mild-moderate weight loss

Numerous experimental and clinical studies have shown that skeletal muscle apoptotis may increase in wasting conditions and suggest that apoptosis might contribute to the loss of lean body mass. Data in cancer patients are still lacking. The present study aimed at verifying whether apoptosis was enhanced in the skeletal muscle of 16 patients with gastric cancer with respect to controls. A biopsy specimen was obtained from the rectus abdominis muscle. The occurrence of apoptosis in muscle biopsies was determined morphologically by the fluorescent transferase-mediated dUTP nick end labeling assay and by immunohistochemistry for caspase-3 and caspase-1. Mean weight loss was 6+/-2% in cancer patients and 0.5+/-0.1% in controls (p<0.0001). Serum albumin levels (g/dL) were 3.7+/-0.3 in cancer patients and 4.1+/-0.2 in controls (p<0.05). The percentage of apoptotic myonuclei was similar in cancer patients and in controls (1.5+/-0.3 versus 1.4+/-0.2, respectively; p=ns), in gastric cancer patients with mild (1.6+/-0.4) or moderate-severe weight loss (1.4+/-0.5) (p=ns), and in the different stages of disease (stages I-II: 1.5+/-0.7; stage III: 1.3+/-0.4; stage IV: 1.6+/-0.3; p=ns). By immunohistochemistry, caspase-1 and caspase-3 positive fibers were absent in controls and in neoplastic patients. Poly-ADP-ribosyl polymerase, a typical caspase-3 substrate whose processing is indicative of caspase-3 activation, was not cleaved in muscle biopsies of cancer patients. These data suggest that skeletal muscle apoptosis is not increased in neoplastic patients with mild-moderate weight loss and argue against the hypotheses that caspase-3 activation might be an essential step of myofibrillar proteolysis in cancer-related muscle wasting.     

Whole rat electron paramagnetic resonance imaging of a nitroxide free radical by a radio frequency (280 MHz) spectrometer.

Low frequency (280 MHz) electron paramagnetic resonance spectroscopy has been used to follow uptake, distribution and reduction of the nitroxyl spin label PCA in the rat. No difference of half life was found in seven rats submitted to three administrations of PCA (11.3 +/- 0.4; 11.0 +/- 0.6 and 11.5 +/- 0.7 min). Transversal two-dimensional images of PCA distribution in the rat body were obtained over 6 min by means of field gradients. PCA was observed in three regions by projections along the longitudinal axis of the rat. PCA accumulation was found in the lower abdomen 12 min after the start of the PCA injection.