The objective of the present experiment was to assess the involvement of small intestine in expression of susceptibility or resistance to the high-fat/high-energy diet. The investigation was carried out in adult male Sprague-Dawley rats fed either standard laboratory diet (3.2 kcal/g, 9.5 % fat) or high-fat (HF) diet (4.04 kcal/g, 30 % fat) for 4 weeks as well as in HF rats that were retrospectively designated on the bases of their higher or lower weight gain as sensitive (DIO) or resistant (DR) to obesity. Our results revealed in HF group significant increase in energy intake, food efficiency, weight gain and Lee´s index of obesity. Moreover, in comparison with controls, a significantly increased duodenal and jejunal alkaline phosphatase (AP) and α-glucosidase activity as well as hypertrophy of jejunal mucosa (increased protein/DNA ratio) were observed in HF fed rats. In contrast, intestinal function was inversely related to energy intake or to the development of adiposity in DIO vs. DR rats. The DR rats had significantly greater AP and α-glucosidase activity and more pronounced suppression of energy intake than obese DIO rats. It indicates that the increase of enzyme activities and the lowered effectiveness of nutrient absorption might be a significant factor preventing the expression of obesity proneness. This information contributes to a better understanding of a complex interaction between HF diet feeding and small intestinal adaptability, which determines the energy homeostasis and predict the ability to resist or develop obesity in these phenotypes., Z. Šefčíková, T. Hájek, Ľ. Lenhardt, Ľ. Raček, Š. Možeš., and Obsahuje bibliografii a bibliografické odkazy
a1_Two experiments were performed to compare the effect of pectin and its hydrophobic derivatives on homeostasis of cholesterol and cecal metabolism in male young rats. Control rats were fed a diet supplemented with palm fat and cholesterol (50 and 10 g/kg, respectively). Rats of other gro ups were fed the same diet containing citrus pectin or octadecylpectinamide (60 g/kg). Diets were fed for 4 weeks. In experiment I, pectinamide of lower degree of amidation (30 %) increased serum HDL cholesterol from 1.20 to 1.43 μmol/ml (p>0.05) at the expense of other cholesterol fractions. In experiment II, pectinamide of a higher degree of amidation (53 %) significantly decreased total serum cholesterol from 2.08 to 1.67 μmol/ml. Amidated pectins at both levels of substitution significantly decreased hepatic concentrations of cholesterol and fat. In both experiments the relative weight of cecum in the pectinamide group was significantly lower than in pectin group. The highest cecal concentrations of short-chain fatty acids (SCFA) were found in rats fed a diet with pectin (133.2 and 129.3 μmol/g in experiment I and II, respectively). In other groups, cecal SCFA was significantly (pectinamide groups) or non-significantly (controls) lower. In wet feces, SCFA concentrations were higher and butyrate molar proportions lower than in corresponding cecal contents., a2_Pectinamide of a lower or higher degree of substitution significantly increased fecal content of cholesterol from 18.5 and 17.3 μmol/g in controls to 31.8 and 28.0 μmol/g, respectively. Corresponding concentrations of coprostanol were decreased. Effects of pectin on cholesterol homeostasis were absent or marginal. Histological examination revealed that hepatic tissue of control and pectin-fed rats was infiltrated with lipids. The Sudan black-positive material was absent in the liver of rats fed pectinamides. No pathological changes of liver tissue were apparent. In summary, hydrophobic amidated pectins significantly altered cholesterol homeostasis in rats and might be considered as a clinically effective hypocholesterolemic agent. Low cecal SCFA concentrations in rats fed pectinamides suggest that amidation of pectin had decreased its fermentability., M. Marounek, Z. Volek, A. Synytsya, J. Čopíková., and Obsahuje bibliografii a bibliografické odkazy
To investigate the relationship between early nutritional experience, ontogeny of the small intestinal functions and predisposition to obesity development, the following experimental models of male Sprague-Dawley rats were used: 1) rats in which the quantity of nutrition was manipulated from birth to weaning (day 30) by adjusting the number of pups in the nest to 4 (SL), 10 (NL) and 16 pups (LL) and 2) littermates of SL, NL and LL rats fed either a standard or a hypercaloric diet from days 80 to 135 of age. The overfed SL pups were overweight after day 15 and became permanently obese, whereas the underfed smaller LL pups, due to accelerated growth and enhanced food intake from day 30 to day 35, attained a body fat level that did not differ from normally fed NL rats. Moreover, a significantly increased duodenal and jejunal alkaline phosphatase (AP) activity was found in SL and LL rats and these acquired somatic and intestinal characteristics persisted from weaning throughout life. Eight weeks of high-energy diet feeding elicited a similar pattern of intestinal response in SL and LL rats that was clearly different from NL rats. Despite energy overconsumption in these three groups, both SL and LL rats still displayed enhanced AP activity and showed a significant increase in protein/DNA ratio accompanied with a significant body fat accretion. These results indicate that the postnatally acquired small intestinal changes induced by over- and undernutrition could be involved in the similar predisposition to obesity risk in later life when caloric density of the diet is raised., Š. Možeš, Z. Šefčíková, Ľ. Lenhardt., and Obsahuje bibliografii a bibliografické odkazy
We have separated 2b myosin heavy chain (MyHC) isoform from the rat extensor digitorum longus muscle by SDS-PAGE and analyzed it by two subsequent mass spectrometry techniques. After tryptic digestion, the obtained peptides were identified by Matrix-Assisted Laser Desorption/Ionisation reflectron Time of Flight mass spectrometry (MALDI-TOF MS) and sequenced by Liquid chromatography tandem mass spectrometry (ESI LC/MS/MS). The analyzed peptides proportionally covered 30 % of the 2b MyHC isoform sequence. The results suggest that the primary structure is identical with the highest probability to a NCBI database record ref|NP_062198.1|, representing the last updated record of rat 2b isoform. Nonetheless, four peptides carrying amino acid substitution(s) in comparison with the NCBI database record were identified., J. Žurmanová, D. Maláčová, F. Půta, P. Novák, J. Říčný, T. Soukup., and Obsahuje bibliografii a bibliografické odkazy
To understand the contribution of potassium (K+) channels, particularly α-dendrotoxin (D-type)-sensitive K+ channels (Kv.1, Kv1.2 or Kv1.6 subunits), to the generation of neuronal spike output we must have detailed information of the functional role of these channels in the neuronal membrane. Conventional intracellular recording methods in current clamp mode were used to identify the role of α-dendrotoxin (α-DTX)-sensitive K+ channel currents in shaping the spike output and modulation of neuronal properties of cerebellar Purkinje neurons (PCs) in slices. Addition of α-DTX revealed that D-type K+ channels play an important role in the shaping of Purkinje neuronal firing behavior. Repetitive firing capability of PCs was increased following exposure to artificial cerebrospinal fluid (aCSF) containing α-DTX, so that in response to the injection of 0.6 nA depolarizing current pulse of 600 ms, the number of action potentials insignificantly increased from 15 in the presence of 4-AP to 29 action potentials per second after application of DTX following pretreatment with 4-AP. These results indicate that D-type K+ channels (Kv.1, Kv1.2 or Kv1.6 subunits) may contribute to the spike frequency adaptation in PCs. Our findings suggest that the activation of voltage-dependent K+ channels (D and A types) markedly affect the firing pattern of PCs., H. Haghdoust, M. Janahmadi, G. Behzadi., and Obsahuje bibliografii a bibliografické odkazy
C-type natriuretic peptides (CNP) play an inhibitory role in smooth muscle motility of the gastrointestinal tract, but the effect of CNP on delayed rectifier potassium currents is still unclear. This study was designed to investigate the effect of CNP on delayed rectifier potassium currents and its mechanism by using conventional whole-cell patch- clamp technique in guinea-pig gastric myocytes isolated by collagenase. CNP significantly inhibited delayed rectifier potassium currents [IK (V)] in dose-dependent manner, and CNP inhibited the peak current elicited by depolarized step pulse to 86.1±1.6 % (n=7, P<0.05), 78.4±2.6 % (n=10, P< 0.01) and 67.7±2.3 % (n=14, P<0.01), at concentrations of 0.01 μmol/l, 0.1 μmol/l and 1 μmol/l, respectively, at +60 mV. When the cells were preincubated with 0.1 μmol/l LY83583, a guanylate cyclase inhibitor, the 1 μmol/l CNP-induced inhibition of IK (V) was significantly impaired but when the cells were preincubated with 0.1 μmol/l zaprinast, a cGMP-sensitive phosphodiesterase inhibitor, the 0.01 μmol/l CNP-induced inhibition of IK (V) was significantly potentiated. 8-Br-cGMP, a membrane permeable cGMP analogue mimicked inhibitory effect of CNP on IK (V). CNP-induced inhibition of IK (V) was completely blocked by KT5823, an inhibitor of cGMP-dependent protein kinase (PKG). The results suggest that CNP inhibites the delayed rectifier potassium currents via cGMP-PKG signal pathway in the gastric antral circular myocytes of the guinea-pig., H. Y. Xu, X. Huang, M. Yang, J.-B. Sun, L.-H. Piao, Y. Zhang, L. Gao, W.-X. Xu., and Obsahuje bibliografii a bibliografické odkazy
The aim of our study was to test the in fluence of short exposure (6 h) of preimplantation rabbit embryos to elevated temperatures (41.5 ºC or 42.5 ºC) in vitro on their developmental capacity. Fertilized eggs recovered from female oviducts at the pronuclear stage (19 hpc) were cultured at standard temperature (37.5 ºC) until the morula stage (72 hpc). Afterwards, the embryos were divided into two groups, cultured for 6 h either at hyperthermic (41.5 ºC or 42.5 ºC) or standard temperature (control 37.5 ºC), post-incubated overnight (16-20 h) at 37.5 ºC and then evaluated for developmental stages, apoptosis (TUNEL), proliferation (cell number), actin cytoskeleton and presence of heat-shock proteins Hsp70. It was observed that hyperthermia at 41.5 ºC did not alter progression of embryos to higher preimplantation stages (expanded and hatching/hatched blastocysts), rate of apoptosis, total cell number of blastocysts and structure of actin filament compared to 37.5 ºC. We stern-blotting revealed the presence of heat stress-induced 72 kDa fraction of Hsp70 proteins in granulosa cells (exposed to 41 ºC) and embryos (exposed to 41.5 ºC). Following the elevation of temperature to 42.5 ºC embryo development was dramati cally compromised. The embryos were arrested at the morula or early blastocyst stage, showed an increased rate of apoptosis and decreased total cell number compared to control. The structure of actin filaments in most of blastomeres was damaged and such blastomeres often contained apoptotic nuclei. In this group a presence of heat-stress-induced fraction of Hsp70 proteins had not been confirmed. This is the first report demonstrating a threshold of thermotolerance of rabbit preimplantation embryos to hyperthermic exposure in vitro. A detrimental effect of higher temperature on the embryo is probably associated with the loss of their ability to produce Hsp70 de novo, which leads to cytoskeleton alterations and enhanced apoptosis., A. V. Makarevich, L. Olexiková, P. Chrenek, E. Kubovičová, K. Fréharová, J. Pivko., and Obsahuje bibliografii a bibliografické odkazy