An attempt was made to assess whether the choice of the gradient media could influence the yield of basolateral membrane vesicles isolated from the rat intestine as well as their functional characteristics. Crude membranes prepared in the same way were therefore centrifuged with 10 % Percoll, on a discontinuous sucrose gradient or on a continuous sorbitol gradient. The protein yield was significantly higher with the Percoll gradient than with sucrose and sorbitol gradient centrifugation (2.7 ±1.0 %; 0.4 ±0.1 %; 0.6±0.2 %, respectively). Enrichment in Ma+,K+-ATPase was similar in all three preparations (8.50±2.34; 8.22±4.78; 8.20±2.08). However, contamination with brush border membranes was significantly higher after Percoll gradient centrifugation and negligible after the use of the other two gradient media. Transport of D-glucose in the BLM prepared by Percoll gradient centrifugation also indicated some contamination with functional brush-border membranes. An attempt to purify basolateral membrane vesicles after Percoll gradient centrifugation with Ca2+ precipitation, however, reduced the protein yield to less than 1 %. We conclude that in the preparation of basolateral membrane vesicles from the rat enterocytes each of the gradient media may have certain advantages and disadvantages, which should be considered according to the purpose of the preparation.
Increasing hemodynamic load during early postnatal development leads to rapid growth of the left ventricular (LV) myocardium, which is associated with membrane phospholipid (PL) remodeling characterized by n-3 polyunsaturated fatty acids (PUFA) accumulation. The aim of this study was to examine the influence of additional workload imposed early after birth when ventricular myocytes are still able to proliferate. Male Wistar rats were subjected to abdominal aortic constriction (AC) at postnatal day 2. Concentrations of PL and their fatty acid (FA) profiles in the LV were analyzed in AC, sham-operated (SO) and intact animals on postnatal days 2 (intact only), 5 and 10. AC resulted in LV enlargement by 22 % and 67 % at days 5 and 10, respectively, compared with age-matched SO littermates. Concentrations of phosphatidylcholine, cardiolipin, phosphatidylinositol, phosphatidylethanolamine, phosphatidylserine and sphingomyelin decreased in AC myocardium, albeit with different time course and extent. The main effect of AC on FA remodeling consisted in the accumulation of n-3 PUFA in PL. The most striking effect of AC on FA composition was observed in phosphatidylinositol and cardiolipin. We conclude that excess workload imposed by AC inhibited the normal postnatal increase of PL concentration while further potentiating the accumulation of n-3 PUFA as an adaptive response of the developing myocardium to accelerated growth., F. Novák, ... [et al.]., and Obsahuje seznam literatury
We have studied in vitro alveolar macrophages (AMs) obtained by tracheobronchial lavage from rats exposed to subacute (3 hours and 3 days) and chronic (3 weeks) hypoxia (Fi02 = 0.1) and from rats recovering from chronic hypoxia. Hydrogen peroxide production by AMs was measured by luminol- depcndent chemiluminescence after AMs adhered to the walls of the measuring cuvette, after stimulation with phorbol-myristate-acetate (PMA), and when N-formyl-methionyl-leucyl-phenylanine (FMLP) was added subsequently to the cells which had been previously stimulated by adherence or PMA. H2O2 production after cell adherence and adherence combined with FMLP stimulation did not differ between the groups. The increase of H2O2 production after adding PMA, and FMLP in addition to PMA was significantly higher in AMs from rats exposed to hypoxia for 3 days than in the controls. Other experimental groups did not differ from their controls. It is concluded that 3 days’ hypoxia primes AMs for enhanced production of H2O2 upon stimulation. The mechanism is probably at the level of synthesis of proteins involved in H2O2 production, or the shift to a more reactive phenotype of alveolar macrophages subpopulations.
The organization of the neocortical projection to the inferior colliculus (IC) was studied in 36 rats using retrograde transport of horseradish peroxidase (HRP) or horseradish peroxidase conjugated with lectin (WGA-HRP). Projection to the external and dorsal cortices originates in the temporal neocortical areas Te 1, Te 2 and Te 3 and in the parietal area Par 2. The corticocollicular projection is predominantly ipsilateral with a weak contralateral contribution. Projection to the rostromedial and rostrolateral part of the external cortex (EC) of the IC arises mainly from the areas Par 2 and Te 1. The participation of the cortical areas Te 2 and Te 3 in this projection is only small. The fibres to the caudobasal part of the external cortex descend from the caudal parts of areas Te 1, Te 2, and Te 3. The corticocollicular projections to the dorsal part of the IC are more numerous than the projections to the EC and originate in all temporal areas, i.e. in area Te 1, Te 2 and Te 3. However, the topographical organization of the corticocollicular projection is more pronounced in the part which projects to the EC. We suggest that the topographical organization of the projections to the EC corresponds with the map of auditory space in the EC. The source of corticocollicular fibres are exclusively neurones of lamina V of all cortical areas sending their fibres to the IC.
Acute lung injury was induced by intravenous injection of 20 //I of a mixture of equivalent volumes of capronic acid, caprilic acid and olive oil in intact anaesthetized rats and in isolated perfused rat lung preparations. Lung injury in intact rats resulted in an increase in lung weight related to body weight and in a decrease in the lung dry/wet weight ratio. Lung compliance, measured in a body plethysmograph, was decreased. Pao2 decreased and Pac02 increased in 10 and 20 min, respectively, after the beginning of the experiment. Mean blood pressure in pulmonary artery increased immediately after the injection. Isolated rat lungs were perfused at constant flow with physiological saline solution containing bovine albumin and meclofenamate. The injection of a mixture of capronic acid, caprilic acid and olive oil increased the baseline perfusion pressure and led to a release of endothelial cells into the perfusate. The perfusion flow-pressure relationship was shifted upwards. Both the extrapolated pressure axis, intercept and slope of the plot were significantly elevated. The described experimental lung injury is a suitable model for studies on the effects of vascular wall damage and transvascular fluid leak in pulmonary vasculature.
An anterograde biocytin and a retrograde WGA-colloidal gold study in the rat can provide information about reciprocal communication pathways between the red nucleus and the trigeminal sensory complex. No terminals were found within the trigeminal motor nucleus, in contrast with the facial motor nucleus. A dense terminal field was observed in the parvicellular reticular formation ventrally to the trigeminal motor nucleus. The parvicellular area may be important for the control of jaw movements by rubrotrigeminal inputs. On the other hand, the contralateral rostral parvicellular part of the red nucleus receives terminals from the same zone in the rostral part of the trigeminal sensory complex, where retrogradely labelled neurones were found after tracer injections into the red nucleus. Such relationships could be part of a control loop for somatosensory information from the orofacial area.
Reciprocal interactions between intralaminar thalamic nuclei (ncl. centralis lateralis, CL, and ncl. parafascicularis, Pf), the pretectal area (Pt) and lateral thalamic nuclei (ventrobasal complex, VB, ncl. anterior ventralis, AV, and ncl. ventralis anterior, VA) have been observed in ketamine-anaesthetized rats. Extracellular single unit activity has been recorded after single electrical stimuli. Electrical stimulation of the VB evoked a short latency orthodromic response followed by a pause in spontaneous activity in neurones of medial thalamic nuclei. Lateral thalamic neurones responded to electrical stimulation of the intralaminar nuclei or the pretectal area with the same pattern of response. Striatal, sensorimotor cortical or peripheral electrical stimulation also evoked similar responses. The pauses in spontaneous activity were shown to be the result of inhibition since the responsiveness of the intralaminar nuclei or the lateral thalamic neurones to all inputs was abolished or reduced after a conditioning electrical singleshock stimulation in the VB or in the intralaminar nuclei, respectively. The two components of the response were of a different origin, since most of the short latency responses disappeared after medullary, upper cervical sections or large decortications, while the inhibitions persisted. These inhibitions were shown to be of thalamic origin since their duration was decreased after extensive decortications increased after medullary section. It is concluded that the neuroneal properties studied in this report are probably broadly represented throughout the thalamus and that thalamic neurones are under inhibitory control elicited by afferent volleys. This inhibitory control includes a relay in the nucleus reticularis thalami (nRT). The mechanisms of sensory interaction can be purely thalamic, but they can be modulated by suprathalamic and/or mesencephalic loops.
The aim of our study was to verify the relationship between heart rate (HR) and ventricular fibrillation threshold (VFT) during different types of ventilation in female Wistar rats from the circadian point of view. The ex-periments were performed under pentobarbital anesthesia (40 mg/kg i.p., adaptation to a light-dark cycle 12:12 h, open chest experiments) and the obtained results were averaged independently of the seasons. The VFT measure-ments were performed during normal ventilation (17 animals) and hypoventilation (10 animals). The HR was re-corded immediately before the rise of ventricular arrhy-thmias. Results are expressed as arithmetic means ± S.D. and differences are considered significant when p<0.05. The basic pe-riodic characteristics were calculated using single and population mean cosinor tests. The results from our experiments have demonstrate that 1) the VFT and HR respond identically to hypoventilation by a decrease in the light and also in the dark phases, and 2) hypoventilation changes the 24-h course of the VFT without a change in the 24-h rhythm of the HR. It is concluded that the HR and VFT behave as two independent functional systems without apparent significant circadian dependence during both types of ventilation., P. Švorc, I. Bračoková, I. Podlubný., and Obsahuje bibliografii
During resetting of the mammalian circadian clock, not only phase of the clock is shifted, but amplitude of overt rhythms driven by the clock may be temporarily reduced or even abolished. The present paper is aimed to elucidate the mechanism of amplitude reduction of the overt circadian rhythm in the rat pineal N-acetyltransferase (NAT). The rhythm has two phase markers, namely the time of the evening NAT rise and that of the morning decline. When the phase relationship between both markers is compressed drastically, the NAT rise may occur just close to or at the time of the decline and consequently the NAT rhythm with a full amplitude cannot be expressed. Such a compression may occur in two ways: either animals are subjected to a considerable advance in the light onset which phase advances the morning NAT decline and at the same time phase delays the evening NAT rise, or they are subjected to a considerable delay in the light offset, which primarily phase delays more the NAT rise than the decline. While in the former case the phase markers move in opposite directions, in the latter case they move in the same direction, but to a different extent. The data suggest a complex structure of the underlying clock.
Ketamine, an N-methyl-D-aspartate antagonist, reduces pain by decreasing central sensitization and pain windup. However, chronic ketamine use can cause tolerance, dependency, impaired consciousness, urinary symptoms, and abdominal pain. This study aimed to investigate the effects of repeated ketamine injections and ketamine readministration after discontinuation in a rat model of neuropathic pain. To induce neuropathic pain, partial sciatic nerve ligation (PSNL) was performed in 15 male Wistar rats, and these animals were divided into three groups: PSNL (control), PSNL + ketamine 5 mg/kg (K5), and PSNL + ketamine 10 mg/kg (K10; n=5 each). Ketamine was injected intraperitoneally daily for 4 weeks, discontinued for 2 weeks, and then readministered for 1 week. Following PSNL, the mechanical withdrawal threshold was determined weekly using the Von Frey. The K10 group showed a significant increase in the mechanical withdrawal threshold, presented here as the target force (in g), at 21 and 28 days compared to the time point before ketamine injection (mean±SE, 276.0±24.0 vs. 21.6±2.7 and 300.0±0.0 vs. 21.6±2.7, respectively; P<0.01) and at 14, 21, and 28 days compared to the control group (108.2±51.2 vs. 2.7±1.3, 276.0±24.0 vs. 2.5±1.5, and 300.0±0.0 vs. 4.0±0.0, respectively; P<0.05). However, in the K10 group, the ketamine effects decreased significantly at 7 days after readministration compared to those after 28 days of repeated injections (P<0.05). In the K10 group, repeated ketamine injections showed a significant increase in antinociceptive effect for >2 weeks, but this ketamine effect decreased after drug readministration.