Glucose and octanoate utilization by isolated adult rat heart cells

Rat heart muscle cells continue to beat in the isolated state apparently independent of any innervation. The oxidation of ¹⁴C-glucose to ¹⁴CO₂ was linear for at least 60 minutes of incubation. The rate of glucose oxidation rose rapidly up to a medium glucose concentration of 2.5 mM and then plateaued. Lactate production reached a maximum at 5 mM glucose. Glucose uptake was linearly related to the concentration up to 40 mM. The addition of octanoate reduced, but did not eliminate, glucose oxidation. Octanoate utilization increased with increasing concentration and reached a maximum at 2 mM. The oxidation of octanoate was linearly related to the time of incubation for at least 90 minutes. The presence of glucose, at a concentration of 1.25 mM or higher, increase the oxidation of octanoate by the heart cells. The metabolic parameters measured with the isolated heart cells gave values comparable to those obtained with the perfused rat heart. Decreasing or increasing the concentration of sodium, potassium or magnesium did not effect the oxidation of either glucose or octanoate with the exception that when sodium was increased above 200 mM, a significant increase in glucose oxidation was observed. In contrast, the addition of calcium to a calcium free medium increased glucose oxidation, reaching a maximum at 0.2 mM calcium. The oxidation of octanoate reached a maximum at 0.2 mM and then decreased significantly with increasing calcium concentration. The metabolic activity appears to be independent of the concentration of sodium, potassium or magnesium. In contrast, the isolated heart cell is very sensitive to a change in calcium concentration.     

Differential effects of fatty acids on glycolysis and glycogen metabolism in vascular smooth muscle

The effects of fatty acids of different chain lengths on aerobic glycolysis, lactic acid production, glycogen metabolism and contractile function of vascular smooth muscle were investigated. Porcine carotid artery segments were treated with 50 microM iodoacetate and perchloric acid tissue extracts were then analyzed by 31P-NMR spectroscopy to observe the accumulation of phosphorylated glycolytic intermediates so that the activity of the Embden-Myerhof pathway could be tracked under various experimental paradigms. Aerobic glycolysis and lactate production in resting arteries were almost completely inhibited with 0.5 mM octanoate, partially inhibited with 0.5 mM acetate and unaffected by 0.5 mM palmitate. Inhibition of glycolysis by octanoate was not attributable to inhibition of glucose uptake or glucose phosphorylation. Basal glycogen synthesis was unchanged with palmitate and acetate, but was inhibited by 52% with octanoate incubation. The characteristic glycogenolysis which occurs upon isometric contraction with 80 mM KCl in the absence of fatty acid in the medium was not demonstrable in the presence of any of the fatty acids tested. Glycogen sparing was also demonstrable in norepinephrine contractions with octanoate and acetate, but not with palmitate. Additionally, norepinephrine-stimulated isometric contraction was associated with enhanced synthesis of glycogen amounting to 6-times the basal rate in medium containing octanoate. Contractile responses to norepinephrine were attenuated by 20% in media containing fatty acids. Thus, fatty acids significantly alter metabolism and contractility of vascular smooth muscle. Fatty acids of different chain lengths affect smooth muscle differentially; the pattern of substrate utilization during contraction depends on the contractile agonist and the fatty acid present in the medium.     

Fatty acids and glycerol or lactate are required to induce gluconeogenesis from alanine in isolated rabbit renal cortical tubules

Summary In isolated rabbit renal cortical tubules, glucose synthesis from 1 mM alanine is negligible, while the amino acid is metabolized to glutamine and glutamate. The addition of 0.5 mM octanoate plus 2 mM glycerol induces incorporation of [U-¹⁴C]Alnine into glucose and decreases glutamine synthesis, whereas oleate and palmitate in the presence of glycerol are less potent than octanoate. Gluconeogenesis is also significantly accelerated when glycerol is substituted by lactate. In view of an increase in¹⁴CO₂ fixation and elevation of both cytosolic and mitochondrial NADH/NAD⁺ ratios, the activation of glucose formation from alanine upon the addition of glycerol and octanoate is likely due to (i) stimulation of pyruvate carboxylation, (ii) increased availability of NADH for glyceraldehyde-3-phosphate dehydrogenase and (iii) elevation of mitochondrial redox state causing a diminished provision of ammonium for glutamine synthesis. The induction of gluconeogenesis in the presence of alanine, glycerol and octanoate is not related to cell volume changes. The results presented in this paper show the importance of free fatty acids and glycerol for regulation of renal gluconeogenesis from alanine. The possible physiological significance of the data is discussed.     

Mitochondrial NAD(P)H, ADP, oxidative phosphorylation, and contraction in isolated heart cells

To examine the relationship between mitochondrial NADH (NADH(m)) and cardiac work output, NADH(m) and the amplitude and frequency of the contractile response of electrically paced rat heart cells were measured at 25 degrees C. With 5.4 mM glucose plus 2 mM beta-hydroxybutyrate, NADH(m) was reversibly decreased by 23%, and the amplitude of contraction was reversibly decreased by 27% during 4-Hz pacing. With glucose plus 2 mM pyruvate or with 10 mM 2-deoxy-D-glucose, NADH(m) was maintained during rapid pacing, and the contractile amplitude remained high. Phosphocreatine levels decreased with 2-deoxy-D-glucose administration but not with rapid pacing. Respiration increased to meet the increased ATP demand at 30 degrees C. The data suggest that 1) when NADH(m) is decreased during rapid pacing with defined substrates, the amplitude of contraction is decreased; 2) the amplitude of contraction during electrical pacing does not change with rate of pacing when both the ATP and NADH(m) levels are continuously replenished; and 3) the replenishment of NADH(m) during pacing with physiological substrates may be rate-limited by substrate supply to mitochondrial dehydrogenases. During activation of mitochondrial dehydrogenases, or a significant increase in free ADP induced by 2-deoxy-D-glucose, this rate limitation is bypassed or overcome.     

cAMP dependence of Ca2+ entry during agonist-induced contractions of the uterine smooth muscles

Possible involvement of cAMP-dependent mechanisms in the development of both phasic and tonic contractions induced by oxytocin — OT (25 nM and 25 µM, respectively), as well as of KCl-induced contracture, was studied on the myometrium of estradiol-dominated rats using the myometrial strips with suppressed spontaneous mechanical activity. The intracellular cAMP level was modulated by furosemide that had been previously shown to decrease cAMP content in the rat myometrium tissue. When added to the medium in the pulse mode together with 25 nM OT, furosemide (0.02 mM) increased contraction amplitude by 224%, whereas higher, 0.2 and 20 mM, furosemide concentrations suppressed the response by 286% or totally removed it, respectively. Being present in the bath permanently, 0.2 mM furosemide progressively decreased the amplitude of OT-induced phasic contractions. Under such conditions, 0.02 mM furosemide exerted biphasic effect on the responses, so that the initial enhancement was replaced by the progressive inhibition. Dibutyryl-cAMP (dbcAMP) at a proper concentration restored the responsiveness of the tissue to OT in the presence of furosemide in the saturating concentration. Contractile responses induced by 25 µM OT comprised both phasic and tonic components. In a Ca²⁺-free medium, the OT-induced contractions seemed to be associated with Ca²⁺ release from intracellular stores. Permanent presence of furosemide in the CaCl₂-containing medium inhibited OT-induced responses in the same manner as omission of Ca²⁺ from the medium, i.e., furosemide did not affect the responses caused by Ca²⁺ release but inhibited those mediated via acceleration of the Ca²⁺ influx. The furosemide-sensitive component of responses to OT was combined with a persistent contraction caused by KCl depolarization; there was a moderate decrease in amplitude of the KCl-induced contracture due to furosemide action. The decrease could be prevented by dbcAMP addition. It is suggested that both voltage-gated and receptor-operated Ca²⁺ entries induced by OT are regulated by cAMP-dependent protein kinases, while Ca²⁺ extrusion into the extracellular space does not depend on the intracellular cAMP.Neirofiziologiya/Neurophysiology, Vol. 26, No. 1, pp. 54–60, January–February, 1994.     

The correlation between the increase in slow outward current and in contraction induced by caffeine, ryanodine, and rapid cooling in voltage-clamped frog muscle fibers.

The effects of caffeine, ryanodine, and rapid cooling were tested on the depolarization-induced contraction and the apamin-insensitive slow outward current (Iso) of voltage-clamped (double mannitol gap) single frog muscle fibers. Subthreshold caffeine concentrations (0.5-2 mM) induced a monotonic increase in contractile and Iso amplitude. Whatever the concentration, the increase in contraction was roughly twice the one in current. Similar results were obtained upon rapid cooling (20-4 degrees C) in the presence of 0.5 mM caffeine. In the absence of external Na+ (choline-substituted) 10(-5) M ryanodine induced a delayed increase (approximately 30 min) in contraction and in current, shortly before the development of a drastic and irreversible contracture. Here again, the increase in contraction was twice that in current. In the presence of 5 mM tetraethylammonium (TEA) and (or) 25 nM charybdotoxin, 2 mM caffeine still induced a strong facilitating effect on contraction but the parallel increase in current was strongly reduced. The linear relationship between the increase in current and contractile amplitude has a slope approximately 0.5 (whatever the drug used to increase contractility); it is approximately 0.1 in the presence of TEA and (or) charybdotoxin. In conclusion, provided the changes in contractile amplitude are caused by parallel changes in depolarization-induced sarcoplasmic reticulum Ca2+ release, about 50% of the apamin-insensitive Iso is controlled by internal Ca2+ release. The main part of this current corresponds to the TEA- and charybdotoxin-sensitive component of Iso.     

Calcium channels and excitation-contraction coupling in cardiac cells. II. A pharmacological study of the biphasic contraction in guinea-pig papillary muscle

Biphasic contractions were obtained in guinea-pig papillary muscle by inducing partial depolarization in K+-rich solution (17 mM) in the presence of 0.3 microM isoproterenol. Mn2+ ions inhibited the two components of contraction in a similar way. Nifedipine and particularly Cd2+ ions specifically inhibited the second component of contraction. Isoproterenol and BAY K 8644 markedly increased the amplitude of the second component (P2) of contraction. Nevertheless, a moderate positive inotropic effect of isoproterenol was found on the first component (P1) of contraction when excitability was restored by 0.2 mM Ba instead of isoproterenol. Acetylcholine and hypoxia decreased the amplitude of the second component of contraction to a greater extent. In the presence of digoxin or Na+-free solution, P1 was strongly increased. When sarcoplasmic reticular function was hindered by 1mM caffeine or in the presence of Ca2+-free Sr2+ solution, digoxin always induced a negative inotropic effect on P2. Inversely in these conditions the transient positive inotropic effect of Na+-free solution was strongly reduced. These results are consistent with the hypothesis that the late component of contraction is triggered by the slow inward Ca2+ current and that the early component is due to Ca2+ release from the sarcoplasmic reticulum.     

Octanoate inhibits very low-density lipoprotein secretion in primary cultures of chicken hepatocytes

The effects of octanoate, a medium-chain fatty acid, on very low-density lipoprotein (VLDL) secretion in primary cultures of chicken hepatocytes were compared with those of palmitate. Palmitate added to the incubation media at concentrations up to 0.36 mM increased intracellular triacylglycerol (TG) accumulation and VLDL-TG secretion in a concentration-dependent manner, whereas the addition of octanoate alone (0.21-0.6 mM) did not change these parameters. VLDL-TG secretion from hepatocytes cultured in media to which 0.6 or 1.0 mM octanoate had been added in the presence of 0.21 mM palmitate was significantly lower than that obtained under control incubation conditions (0.21 mM palmitate only). The addition of 1.0 mM octanoate to the incubation media with or without 0.21 mM palmitate decreased VLDL apolipoprotein B (apoB) secretion. These results demonstrate that the addition of octanoate to primary cultures of chicken hepatocytes reduces VLDL secretion in respect of both TG and apoB secretion. It is suggested that medium-chain fatty acids are a factor modulating VLDL secretion, which plays a key role in fat deposition in chickens.     

Calcium channels and excitation-contraction coupling in cardiac cells. I. Two components of contraction in guinea-pig papillary muscle

Biphasic contractions have been obtained in guinea-pig papillary muscle by inducing partial depolarization in K+-rich solution (17 mM) containing 0.3 microM isoproterenol; whereas in guinea-pig atria, the same conditions led to monophasic contractions corresponding to the first component of contraction in papillary muscle. The relationships between the amplitude of the two components of the biphasic contraction and the resting membrane potential were sigmoidal curves. The first component of contraction was inactivated for membrane potentials less positive than those for the second component. In Na+-low solution (25 mM), biphasic contraction became monophasic subsequent to the loss of the second component, but tetraethylammonium unmasked the second component of contraction. The relationship between the amplitude of the first component of contraction and the logarithm of extracellular Ca2+ concentration was complex, whereas for the second component it was linear. When Ca2+ ions were replaced by Sr2+ ions, only the second component of contraction was observed. It is suggested that the first component of contraction may be triggered by a Ca2+ release from sarcoplasmic reticulum, induced by the fast inward Ca2+ current and (or) by the depolarization. The second component of contraction may be due to a direct activation of contractile proteins by Ca2+ entering the cell along with the slow inward Ca2+ current and diffusing through the sarcoplasm. These results do not exclude the existence of a third "tonic" component, which could possibly be mixed with the second component of contraction.     

Proctolin and octopamine actions on the contractile systems of insect leg muscles

1. The pentapeptide proctolin produced contractions of the coxal depressor muscle of the cockroach, Periplaneta americana.2. The contraction was dependent upon extracellular calcium and the contraction was completely blocked by a Ca-free EGTA saline.3. Caffeine elicited transient contractions which were unaffected by manganese treatment.4. When the muscle was pre-treated with the conditioning solution with different K⁺ concentrations (1–100 mM), the amplitude of proctolin-induced contractions was reduced in the low K⁺ saline as well as in the high K⁺ saline.5. The results suggest that voltage sensitive calcium channels account for the proctolin-induced contractions.6. Octopamine (OA) reduced the contractions resulting from brief applications of elevated K⁺ concentration and of caffeine.7. The effect of OA on the response to elevated K⁺ concentrations was blocked by the α-adrenergic blocker, phentolamine.     

Influence of ryanodine receptor agonist and antagonist on development of potassium contracture in phasic (twitch) and tonic fibers of frog skeletal muscle

We compared the influence of external calcium and the inhibitor (dantrolene) and activator (4-chloro-m-cresol) of ryanodine-sensitive Ca channels of the sarcoplasmic reticulum on the characteristics of potassium contracture in phasic and tonic frog skeletal muscle fibers. The duration of contracture in tonic fibers, as contrasted to the phasic ones, is not limited by the presence of Ca²⁺. The tonic contractile response is virtually indifferent to dantrolene and is much less sensitive to chlorocresol than the phasic one (1 mM vs. 0.25 mM). In phasic fibers, the K⁺ contracture on the chlorocresol background is quite similar in amplitude and dynamics to that in control, whereas tonic fibers exhibit response summation without relaxation upon removal of excessive K⁺. One can suggest that in phasic fibers the Ca²⁺ influx can directly create a level sufficient to sustain contraction, while in tonic fibers its effect is mediated by Ca-dependent activation of the beta isoform of the ryanodine-sensitive channel.     

Modification of efferent vagal effects on the isolated atrium by increased extracellular K+ concentrations

Isolated vagal-innervated rabbit atria are electrically driven. Alterations of action potential, contraction, and electrotropic and inotropic vagal effects are investigated during variations of the external potassium concentration. Action potential area and contraction amplitude decrease by increasing external potassium concentration. If the potassium concentration is higher than 11 mM, the action potential disappears. At 24 mM potassium concentration the contraction amplitude of the driven atrium is reduced to 2%. Adrenaline (2.10(-5) g/ml) causes a restitution of the action potential and the contraction. With increasing potassium concentration the inotropic and the electrotropic vagal effectivity increases also. The vagal effects at the adrenaline restituted action potentials and contractions (15 mM potassium, 2.10(-5) g/ml adrenaline) are also higher than in normal solutions. The relations of electromechanical coupling are altered by potassium variation at the same coupling curve. With increasing potassium concentration the reproducibility of the vagal effects decreases.     

Mechanism of increased conversion of branched chain keto acid dehydrogenase from inactive to active form by a medium chain fatty acid (octanoate) in skeletal muscle.

We and others have previously shown that octanoate increases the oxidation of branched chain amino acids (BCAA) in skeletal muscle. The present study was designed to investigate the mechanism of this increased oxidation. Studies were performed with rat hind limbs perfused with 0.50 mM L-[1-14C]leucine with or without octanoate. The flux through branched chain keto acid (BCKA) dehydrogenase was measured, and the basal and total activity of BCKA dehydrogenase in skeletal muscle was determined. The rate of flux through BCKA dehydrogenase increased by 37, 119, and 297% with 0.5, 1.0, and 2.0 mM octanoate, respectively. This increase in flux was not due to a change in BCAA aminotransferase activity but was due to an increase in the basal activity of BCKA dehydrogenase. There was a strong correlation (r = 0.96) between increases in flux through BCKA dehydrogenase and increases in the basal activities of BCKA dehydrogenase. Preincubation of BCKA dehydrogenase with Mg2+ caused full activation of this enzyme, but preincubation with octanoate did not activate this enzyme. On the other hand, octanoate completely prevented the ATP-dependent inactivation of fully activated BCKA dehydrogenase. We conclude that octanoate increases the oxidation of leucine in skeletal muscle by increasing the activation of BCKA dehydrogenase. The mechanism of this activation is the inhibition of BCKA dehydrogenase kinase rather than the stimulation of a specific or nonspecific protein phosphatase.     

Modification of swelling–contraction–aggregation processes in rat muscle mitochondria by the 1,4-dihydropyridines,cerebrocrast and glutapyrone,themselves and in the presence of azidothymidine

The influence of the 1,4-dihydropyridines (DHPs), water-soluble glutapyrone available as sodium, potassium and ammonium salts of 2-(2,6-dimethyl-3,5-diethoxycarbonyl-1,4-DHP-4-carboxamide)glutaric acid, from one side, and a lipophylic cerebrocrast, 2-propoxyethyl 2,6-dimethyl-4-(2-difluoromethoxyphenyl)-1,4-DHP-3,5-dicarboxylate, from the other side, on partially damaged mitochondria of the Wistar rat hindlimb muscle was also studied. The following tests were made: (1) rates of endogenous respiration and substrate (succinate) oxidation and oxidative phosphorylation; (2) rates and amplitudes of high-amplitude swelling and contraction after the addition of ATP, ADP and succinate to the previously swollen mitochondria and (3) rate of reversible self-aggregation of mitochondria isolated in salt media after ATP-induced contraction without and in the presence of azidothymidine (AZT). Cerebrocrast (10–100 μM ) partially normalized the endogenous respiration rate and slightly augmented the respiration rate after the addition of succinate and to lesser extent ADP. Cerebrocrast in a concentration-dependent manner (2·5–50 μM ) increased (two-fold at 20–50 μM ) the active contraction amplitude of swollen mitochondria, induced by single or repeated additions of ATP. The influence of cerebrocrast on the ADP- and succinate-induced contractions was less obvious. Unlike cerebrocrast glutapyrone caused a reduction of the ATP-induced contraction amplitude (two-fold at 0·5–5·0 mM ), not impairing the mitochondrial contraction ability in response to ATP or succinate. Pre-exposure to 2·5 mM glutapyrone resulted in at least a 10-fold inhibition of the reversible aggregation rate in the presence of 99 and 198 μM AZT. The results suggest the usefulness of further study of cerebrocrast and glutapyrone in preventing AZT-induced and some other mitochondrial myopathies. © 1997 John Wiley & Sons, Ltd.     

The actions of intermediate and long-chain n-alkanols on unitary NMDA currents in hippocampal neurons.

The actions of the n-alkanols butanol, pentanol, and octanol on unitary currents passing through N-methyl-D-aspartate (NMDA) ion channels have been studied in cultured CA1 hippocampal neurons. The cell-attached patch clamp method, with L-homocysteic acid included in the patch pipette, was used to record single channel NMDA currents at the cell resting potential or for hyperpolarizing patch potentials. With the n-alkanols added to the bath solution, the mean open times for the NMDA channel were diminished and the channel conductance was unchanged. A decrease in mean open time to about 70% of control value was found with butanol (3 mM), pentanol (1 mM), and octanol (0.02 mM). In addition the n-alkanols had small effects to decrease the frequency of channel openings and to increase the amplitude of the unitary currents. The effects of the alcohols on intracellular calcium levels, during NMDA applications, were also measured using the fluorescent dye FURA II.     

T-channels and Na<Superscript>+</Superscript>,Ca<Superscript>2+</Superscript>-exchangers as components of the Ca<Superscript>2+</Superscript>-system of regulation of activity of the heart myocardium of the frog <Emphasis Type="Italic">Rana temporaria</Emphasis>

Earlier we have shown that regulation of rhythm and strength of the frog heart contractions, mediated by transmitters of the autonomic nervous system, is of the Ca²⁺-dependent character. In the present work, we studied chronoand inotropic effect of verapamil—an inhibitor of Ca²⁺-channels of the L-type, of nickel chloride-an inhibitor of Ca²⁺—channels of the T-type and of Na⁺,Ca²⁺exchangers as well as of adrenaline and acetylcholine (ACh) after nickel chloride. It has been found that the intracardially administered NiCh₂ at a dose of 0.01 μg/kg produced a sharp fall of amplitude of action potential (AP) and an almost twofold deceleration of heart rate (HR). The intracardiac administration of NiCh₂ (0.01 μg/kg) on the background of action of verapamil (6 mg/kg, i/m) led as soon as after 3 min to even more prominent HR deceleration and to further fall of the AP amplitude by more than 50% as compared with norm. An intracardiac administration of adrenaline (0.5 mg/kg) partly restored the cardiac activity. However, preservation of the myocardium electrical activity in such animals was brief and its duration did not exceed several minutes. Administration of Ni²⁺ on the background of acetylcholine (3.6 mg/kg) led to almost complete cessation of cardiac activity. As soon as 3 min after injection of this agent the HR decreased to 2 contractions/min. On electrograms (EG), the 10-fold fall of the AP amplitude was recorded. To elucidate role of extraand intracellular Ca²⁺ in regulation of strength of heart contractions, isometric contraction of myocardium preparations was studied in response to action of NiCl₂ (10–200 μM), verapamil (70 μM), adrenaline (5 μM), and acetylcholine (0.2 μM) after NiCl₂. It has been found that Ni²⁺ causes a dose-dependent increase of the muscle contraction amplitude. Minimal change of the contraction amplitude (on average, by 14.9% as compared with control) was recorded at a Ni²⁺ concentration of 100 μM. An increase of Ni²⁺ in the sample to 200 μM increased the cardiac contraction strength, on average, by 41%. The negative inotropic action of verapamil was essentially reduced by 100 μM Ni²⁺. Adrenaline added to the sample after Ni²⁺ produced stimulating effect on the cardiac muscle, with an almost twofold rise of the contraction amplitude. ACh (0.2 μM) decreased the cardiac contraction amplitude, on average, by 56.3%, whereas Ni²⁺ (200 μM) administered after ACh not only restored, but also stimulated partly the myocardial work. Within several parts of percent there was an increase of such isometric contraction parameters as amplitude of the effort developed by muscle, maximal rate, maximal acceleration, time of semirise and semifall. The obtained experimental results indicate that the functional activity of the frog pacemaker and contractile cardiomyocytes is regulated by Ca²⁺-dependent mechanisms. Structure of these mechanisms includes the potential-controlled Land T-channels of the plasma membrane as well as Na⁺,Ca²-exchangers characteristic exclusively of contractile cardiomyocytes. The existence of these differences seems to be due to the cardiomyocyte morphological peculiarities that appeared in evolution at the stage of the functional cell specialization.     

Ketogenesis evaluation in perfused liver of diabetic rats submitted to short‐term insulin‐induced hypoglycemia

Ketogenesis, inferred by the production of acetoacetate plus ß‐hydroxybutyrate, in isolated perfused livers from 24‐h fasted diabetic rats submitted to short‐term insulin‐induced hypoglycemia (IIH) was investigated. For this purpose, alloxan‐diabetic rats that received intraperitoneal regular insulin (IIH group) or saline (COG group) injection were compared. An additional group of diabetic rats which received oral glucose (gavage) (100 mg kg^?1) 15 min after insulin administration (IIH + glucose group) was included. The studies were performed 30 min after insulin (1.0 U kg^?1) or saline injection. The ketogenesis before octanoate infusion was diminished (p < 0.05) in livers from rats which received insulin (COG vs. IIH group) or insulin plus glucose (COG vs. IIH + glucose group). However, the liver ketogenic capacity during the infusion of octanoate (0.3 mM) was maintained (COG vs. IIH group and COG vs. IIH + glucose group). In addition, the blood concentration of ketone bodies was not influenced by the administration of insulin or insulin plus glucose. Taken together, the results showed that inspite the fact that insulin and glucose inhibits ketogenesis, livers from diabetic rats submitted to short‐term IIH which received insulin or insulin plus glucose showed maintained capacity to produce acetoacetate and ß‐hydroxybutyrate from octanoate. Copyright © 2009 John Wiley & Sons, Ltd.     

Regulation of myocardial fatty acid oxidation by substrate supply

We tested the hypothesis that myocardial substrate supply regulates fatty acid oxidation independent of changes in acetyl-CoA carboxylase (ACC) and 5'-AMP-activated protein kinase (AMPK) activities. Fatty acid oxidation was measured in isolated working rat hearts exposed to different concentrations of exogenous long-chain (0.4 or 1.2 mM palmitate) or medium-chain (0.6 or 2.4 mM octanoate) fatty acids. Fatty acid oxidation was increased with increasing exogenous substrate concentration in both palmitate and octanoate groups. Malonyl-CoA content only rose as acetyl-CoA supply from octanoate oxidation increased. The increases in octanoate oxidation and malonyl-CoA content were independent of changes in ACC and AMPK activity, except that ACC activity increased with very high acetyl-CoA supply levels. Our data suggest that myocardial substrate supply is the primary mechanism responsible for alterations in fatty acid oxidation rates under nonstressful conditions and when substrates are present at physiological concentrations. More extreme variations in substrate supply lead to changes in fatty acid oxidation by the additional involvement of intracellular regulatory pathways.     

Cellular mechanisms involved in iso-osmotic high K+ solutions-induced contraction of the estrogen-primed rat myometrium

The aim of the present study was to investigate the mechanisms involved in the contraction evoked by iso-osmotic high K+ solutions in the estrogen-primed rat uterus. In Ca2+-containing solution, iso-osmotic addition of KCl (30, 60 or 90 mM K+) induced a rapid, phasic contraction followed by a prolonged sustained plateau (tonic component) of smaller amplitude. The KCl (60 mM)-induced contraction was unaffected by tetrodotoxin (3 microM), omega-conotoxin MVIIC (1 microM), GF 109203X (1 microM) or calphostin C (3 microM) but was markedly reduced by tissue treatment with neomycin (1 mM), mepacrine (10 microM) or U-73122 (10 microM). Nifedipine (0.01-0.1 microM) was significantly more effective as an inhibitor of the tonic component than of the phasic component. After 60 min incubation in Ca2+-free solution containing 3 mM EGTA, iso-osmotic KCl did not cause any increase in tension but potentiated contractions evoked by oxytocin (1 microM), sodium orthovanadate (160 micrM) or okadaic acid (20 microM) in these experimental conditions. In freshly dispersed myometrial cells maintained in Ca2+-containing solution and loaded with indo 1, iso-osmotic KCl (60 mM) caused a biphasic increase in the intracellular Ca2+ concentration ([Ca2+]i). In cells superfused for 60 min in Ca2+-free solution containing EGTA (1 mM), KCl did not increase [Ca2+]i. In Ca2+-containing solution, KCl (60 mM) produced a 76.0 +/- 16.2% increase in total [3H]inositol phosphates above basal levels and increased the intracellular levels of free arachidonic acid. These results suggest that, in the estrogen-primed rat uterus, iso-osmotic high K+ solutions, in addition to their well known effect on Ca2+ influx, activate other cellular processes leading to an increase in the Ca2+ sensitivity of the contractile machinery by a mechanism independent of extracellular Ca2+.