East-to-west human dispersal into Europe 1.4 million years ago.

Stone tools stratified in alluvium and loess at Korolevo, western Ukraine, have been studied by several research groups1-3 since the discovery of the site in the 1970s. Although Korolevo's importance to the European Palaeolithic is widely acknowledged, age constraints on the lowermost lithic artefacts have yet to be determined conclusively. Here, using two methods of burial dating with cosmogenic nuclides4,5, we report ages of 1.42 ± 0.10 million years and 1.42 ± 0.28 million years for the sedimentary unit that contains Mode-1-type lithic artefacts. Korolevo represents, to our knowledge, the earliest securely dated hominin presence in Europe, and bridges the spatial and temporal gap between the Caucasus (around 1.85-1.78 million years ago)6 and southwestern Europe (around 1.2-1.1 million years ago)7,8. Our findings advance the hypothesis that Europe was colonized from the east, and our analysis of habitat suitability9 suggests that early hominins exploited warm interglacial periods to disperse into higher latitudes and relatively continental sites-such as Korolevo-well before the Middle Pleistocene Transition.

A fundamental evaluation of the electrical properties and function of cardiac transverse tubules.

This work discusses active and passive electrical properties of transverse (T-)tubules in ventricular cardiomyocytes to understand the physiological roles of T-tubules. T-tubules are invaginations of the lateral membrane that provide a large surface for calcium-handling proteins to facilitate sarcomere shortening. Higher heart rates correlate with higher T-tubular densities in mammalian ventricular cardiomyocytes. We assess ion dynamics in T-tubules and the effects of sodium current in T-tubules on the extracellular potential, which leads to a partial reduction of the sodium current in deep segments of a T-tubule. We moreover reflect on the impact of T-tubules on macroscopic conduction velocity, integrating fundamental principles of action potential propagation and conduction. We also theoretically assess how the conduction velocity is affected by different T-tubular sodium current densities. Lastly, we critically assess literature on ion channel expression to determine whether action potentials can be initiated in T-tubules.

Metabolic cardio- and reno-protective effects of empagliflozin in a prediabetic rat model.

The mechanisms behind the cardiovascular and renal benefits of empagliflozin is not fully understood. The positive impact of the medication on cardiovascular mortality can not be solely attributed to its antidiabetic effect, with a metabolic mechanism possibly involved. To investigate the metabolic effects of empagliflozin treatment (10 mg/kg/day for 6 weeks), we used an adult male rat model with serious vascular complications associated with metabolic syndrome and prediabetes. Impaired glucose tolerance, severe albuminuria and impaired insulin sensitivity were induced by intragastric administration of methylglyoxal and high sucrose diet feeding for four months. Although empagliflozin decreased body weight, non-fasting glucose and insulin, glucagon levels remained unchanged. In addition, empagliflozin increased adiponectin levels (+40%; p < 0.01) and improved skeletal muscle insulin sensitivity. Increased non-esterified fatty acids (NEFA) in empagliflozin-treated rats is understood to generate ketone bodies. Empagliflozin increased ß-hydroxybutyrate levels in serum (+66%; p < 0.05) and the myocardium (30%; p < 0.01), suggesting its possible involvement as an alternative substrate for metabolism. Empagliflozin switched substrate utilisation in the myocardium, diverting glucose oxidation to fatty acid oxidation. Representing another favorable effect, empagliflozin also contributed to decreased uric acid plasma levels (-19%; p < 0.05). In the kidney cortex, empagliflozin improved oxidative and dicarbonyl stress parameters and increased gene expression of ß-hydroxybutyrate dehydrogenase, an enzyme involved in ketone body utilisation. In addition, empagliflozin decreased microalbuminuria (-27%; p < 0.01) and urinary neutrophil gelatinase-associated lipocalin (NGAL) excretion (-29%; p < 0.01). Our results reveal the important systemic metabolic effect of empagliflozin on alterations in substrate utilisation and on increased ketone body use in prediabetic rats. Improved oxidative and dicarbonyl stress and decreased uric acid are also possibly involved in the cardio- and reno-protective effects of empagliflozin.

Differential sensitivity of skeletal and fusimotor neurons to Bcl-2-mediated apoptosis during neuromuscular development.

Proper development of the nervous system requires that a carefully controlled balance be maintained between both proliferation and neuronal survival. The process of programmed cell death is believed to play a key role in regulating levels of neuronal survival, in large part through the action of antiapoptotic proteins, such as Bcl-2. Consistent with this, Bcl-2 has been shown to be a key regulator of apoptotic signaling in post-mitotic neurons. However, we still know remarkably little regarding the role that Bcl-2 plays in regulating the survival of specific motor neuron populations. In the present study, we have examined somatic motor neurons of the lumbar spinal cord, and branchiomotor neurons of the facial nucleus in bcl-2-null mice to determine the differential dependence among motor neuron populations with respect to Bcl-2-mediated survival. Examination of neuronal and axon number, axonal area, and the distribution of axonal loss in bcl-2-null mice demonstrates that, in contrast to the great majority of alpha motor neurons, gamma motor neurons exhibit a unique dependence upon bcl-2 for survival. These results demonstrate, for the first time, the connection between Bcl-2 expression, motor neuron survival, and the establishment of different motor populations.

Paradoxical improvement of impulse conduction in cardiac tissue by partial cellular uncoupling.

Generally, impulse propagation in cardiac tissue is assumed to be impaired by a reduction of intercellular electrical coupling or by the presence of structural discontinuities. Contrary to this notion, the spatially uniform reduction of electrical coupling induced successful conduction in discontinuous cardiac tissue structures exhibiting unidirectional conduction block. This seemingly paradoxical finding can be explained by a nonsymmetric effect of uncoupling on the current source and the current sink in the preparations used. It suggests that partial cellular uncoupling might prevent the initiation of cardiac arrhythmias that are dependent on the presence of unidirectional conduction block.

Introduction of a neurotrophin-3 transgene into muscle selectively rescues proprioceptive neurons in mice lacking endogenous neurotrophin-3.

To clarify the role of muscle-derived neurotrophin-3 (NT-3) in the development of sensory neurons, we generated transgenic mice selectively overexpressing NT-3 in skeletal muscles under the control of a myogenin promoter (myo-NT-3 mice). The myo-NT-3 transgene was then bred into an NT-3 null mutant (-/-) line to generate myo-NT-3, NT-3(-/-) mice in which NT-3 was expressed in muscles, but not elsewhere. Transient overexpression of NT-3 in developing muscles increased the number of proprioceptive neurons as well as the density of both their central and peripheral projections, resulting in more Ia afferents in spinal cord and more spindles (end organs of Ia afferents) in muscles. NT-3 expression restricted to muscles was sufficient to secure the development of proprioceptive neurons and their central and peripheral projections in myo-NT-3, NT-3(-/-) mice. The loss of nonproprioceptive neurons observed in NT-3(-/-) mice was not reversed by the transgene, suggesting that these neurons are regulated by NT-3 from sources other than muscle. We conclude that target-derived rather than intraganglionic NT-3 is preeminent in supporting the development of proprioceptive neurons. The level of NT-3 in developing muscles may be the principal factor determining the number of proprioceptive neurons in dorsal root ganglions and spindles in skeletal muscles of adults.

Development of sensory neurons in the absence of NGF/TrkA signaling in vivo.

The neurotrophin survival dependence of peripheral neurons in vitro is regulated by the proapoptotic BCL-2 homolog BAX. To study peripheral neuron development in the absence of neurotrophin signaling, we have generated mice that are double null for BAX and nerve growth factor (NGF), and BAX and the NGF receptor TrkA. All dorsal root ganglion (DRG) neurons that normally die in the absence of NGF/TrkA signaling survive if BAX is also eliminated. These neurons extend axons through the dorsal roots and collateral branches into the dorsal horn. In contrast, superficial cutaneous innervation is absent. Furthermore, rescued sensory neurons fail to express biochemical markers characteristic of the nociceptive phenotype. These findings establish that NGF/TrkA signaling regulates peripheral target field innervation and is required for the full phenotypic differentiation of sensory neurons.

Activation of electrical triggers of atrial fibrillation in hyperthyroidism.

CONTEXT: A shortening of the atrial refractory period has been considered as the main mechanism for the increased risk of atrial fibrillation in hyperthyroidism. However, other important factors may be involved. OBJECTIVE: Our objective was to determine the activity of abnormal supraventricular electrical depolarizations in response to elevated thyroid hormones in patients without structural heart disease. PATIENTS AND DESIGN: Twenty-eight patients (25 females, three males, mean age 43+/-11 yr) with newly diagnosed and untreated hyperthyroidism were enrolled in a prospective trial after exclusion of heart disease. Patients were followed up for 16 +/- 6 months and studied at baseline and 6 months after normalization of serum TSH levels. MAIN OUTCOME MEASURES: The incidence of abnormal premature supraventricular depolarizations (SVPD) and the number of episodes of supraventricular tachycardia was defined as primary outcome measurements before the start of the study. In addition, heart rate oscillations (turbulence) after premature depolarizations and heart rate variability were compared at baseline and follow-up. RESULTS: SVPDs decreased from 59 +/- 29 to 21 +/- 8 per 24 h (P = 0.003), very early SVPDs (so called P on T) decreased from 36 +/- 24 to 3 +/- 1 per 24 h (P < 0.0001), respectively, and nonsustained supraventricular tachycardias decreased from 22 +/- 11 to 0.5 +/- 0.2 per 24 h (P = 0.01) after normalization of serum thyrotropin levels. The hyperthyroid phase was characterized by an increased heart rate (93 +/- 14 vs. 79 +/- 8 beats/min, P < 0.0001) and a decreased turbulence slope (3.6 vs. 9.2, P = 0.003), consistent with decreased vagal tone. This was confirmed by a significant decrease of heart rate variability. CONCLUSION: Hyperthyroidism is associated with an increased supraventricular ectopic activity in patients with normal hearts. The activation of these arrhythmogenic foci by elevated thyroid hormones may be an important causal link between hyperthyroidism and atrial fibrillation.

Intraspecific variation in Viola suavis in Europe: parallel evolution of white-flowered morphotypes.

BACKGROUND AND AIMS: Viola species are commonly grown for their ornamental flowers, but their evolutionary history and taxonomy are often complicated and have been poorly explored so far. This is a study of the polymorphic, typically blue-flowered species Viola suavis, concentrating on the white-flowered populations of uncertain taxonomic assignment that occur in Spain and central and south-eastern Europe. The aim was to resolve their origin and taxonomic status and to study the intraspecific structure and (post)glacial history of this species. METHODS: Viola suavis and five close relatives were sampled from multiple locations and subjected to molecular (AFLP, sequencing of nrDNA ITS) and morphometric analyses. Data on ploidy level and pollen fertility were also obtained, to address an assumed hybrid origin of the white-flowered populations. KEY RESULTS: In V. suavis a strong intraspecific genetic split into two groups was observed, indicating that there has been a long-term isolation and survival in distinct glacial refugia. The white-flowered populations could be placed within the variation range of this species, and it is clear that they evolved independently in two distant areas. Their parallel evolution is supported by both morphological and genetic differentiation. The strongly reduced genetic variation and absence of unique AFLP fragments suggest their derived status and origin from the typical, blue-flowered populations. CONCLUSIONS: These results suggest that intraspecific variation in V. suavis has been largely shaped by population isolations during the last glaciation and subsequent recolonizations, although cultivation and vegetative spread by humans have affected the present picture as well.

Mutilating electrotrauma--case report.

The passage of electric current through a human body causes polarization changes in cell membranes, which can possibly lead to the death of these cells. At the same time, electric energy is transformed to thermal energy, primarily in high resistance tissues. We present a case report of a 22-year-old male who was hit by an electric current with a voltage of 22 kV when he was working on a high-voltage overhead line tower. Primary treatment which included fasciotomies was completed two hours after the injury. Fasciotomies and revisions of all muscle groups were completed on the left upper extremity and right shank. On the right upper extremity fasciotomies were completed on the forearm. Retinaculum flexorum was cut in the area of both wrists. Despite the complex therapy including higher doses of a low-molecular-weight heparin, ischemization of the whole left upper extremity and distal part of right shank and foot occurred. On the sixth day after the injury it was necessary to amputate the right lower extremity in shank and on the eighth day after injury to amputate the left upper extremity below the shoulder, and on the fourteenth day, due to progressive ischemic necrosis, it was necessary to complete exarticulation of the left shoulder. The 45th day after the injury our team of micro-surgeons closed the defect of soft tissues in the distal part of right forearm and radial part of right hand by transferred parascapular fasciocutaneous flap. The right median nerve appeared to be necrotic in the distal part of forearm even at the day of injury. Four months after the injury the 12 cm long defect of the right median nerve was bypassed by a graft from the suralis nerve. Outpatient care followed as well as physical and psychological rehabilitation. The support of the family was admirable. One and a half years after the injury reconstruction of the right thumb flexor tendon was completed. Two years after the injury function of the right hand in terms of grip function was satisfactory (patient was able to complete pinch grip and sign). Gait with the prosthesis was very good.

Determination of deuterium oxide content in water based on luminescence quenching.

Water molecules (H2O) often reduce luminescence lifetimes of various luminescence probes. The change of lifetime is usually caused by dynamic luminescence quenching induced by O-H oscillators which effectively take away energy from excited molecule. The process can be described by Stern-Volmer equation. We have studied selected luminescence systems where it is possible to detect considerable changes of lifetime in presence/absence of H2O and D2O in this work for analytical purposes. We have tested both, inorganic (Ln3+) and organic compounds using three different instrumentation in order to find the largest change between τH and τD. The Ln3+ containing systems have shown considerable increase/decrease of lifetimes in the presence/absence of D2O (Eu3+: τD/τH = 34.5) whereas organic systems gave significantly lower values of τD/τH (coumarin 123 lifetime ratio, τD/τH = 1.94). The calculated LOD varied from 0.04 mol l-1 (samarium nitrate) to 6.55 mol l-1 (riboflavin).

Uniaxial strain of cultured mouse and rat cardiomyocyte strands slows conduction more when its axis is parallel to impulse propagation than when it is perpendicular.

AIM: Cardiac tissue deformation can modify tissue resistance, membrane capacitance and ion currents and hence cause arrhythmogenic slow conduction. Our aim was to investigate whether uniaxial strain causes different changes in conduction velocity (θ) when the principal strain axis is parallel vs perpendicular to impulse propagation. METHODS: Cardiomyocyte strands were cultured on stretchable custom microelectrode arrays, and θ was determined during steady-state pacing. Uniaxial strain (5%) with principal axis parallel (orthodromic) or perpendicular (paradromic) to propagation was applied for 1 minute and controlled by imaging a grid of markers. The results were analysed in terms of cable theory. RESULTS: Both types of strain induced immediate changes of θ upon application and release. In material coordinates, orthodromic strain decreased θ significantly more (P < .001) than paradromic strain (2.2 ± 0.5% vs 1.0 ± 0.2% in n = 8 mouse cardiomyocyte cultures, 2.3 ± 0.4% vs 0.9 ± 0.5% in n = 4 rat cardiomyocyte cultures, respectively). The larger effect of orthodromic strain can be explained by the increase in axial myoplasmic resistance, which is not altered by paradromic strain. Thus, changes in tissue resistance substantially contributed to the changes of θ during strain, in addition to other influences (eg stretch-activated channels). Besides these immediate effects, the application of strain also consistently initiated a slow progressive decrease in θ and a slow recovery of θ upon release. CONCLUSION: Changes in cardiac conduction velocity caused by acute stretch do not only depend on the magnitude of strain but also on its orientation relative to impulse propagation. This dependence is due to different effects on tissue resistance.

Experience in non-conventional wastewater treatment techniques used in the Czech Republic.

Among the most common non-conventional wastewater treatment techniques used in the Czech Republic are waste stabilisation ponds (WSP), subsurface horizontal flow constructed wetlands (CW) and vertical flow groundfilters (GF). These extensive systems can be advantageously used for treatment of waters coming from sewerages where the ballast weighting commonly makes more than half of dry-weather flow. The monitoring was focused at 14 different extensive systems. Organics removal efficiencies were favourable (CW-82%; GF-88%); in the case of WSP only 57% due to the algal bloom. Total nitrogen removal efficiencies were 43 and 47% for WSP and GF; in the case of CW only 32% due to often occurring anaerobic conditions in filter beds. Total phosphorus removal efficiencies were 37, 35 and 22% for WSP, GF and CW, respectively. Often occurring problems are the ice-blockage of surface aerators at WSP during wintertimes, the pond duckweed-cover or the algal bloom at WSP during summers; a gradual colmatage of filter systems; and the oxygen deficiency in beds of subsurface horizontal flow constructed wetlands. Czech legal regulations do not allow treated wastewater disposal into underground waters. There is only an exception for individual family houses. Up to now, knowledge gained by monitoring of a village (which uses the infiltration upon a permission issued according to earlier legal regulations) have not shown an unacceptable groundwater quality deterioration into the infiltration areas.

[Approximative intestinal anastomosis in newborns with multifocal necrotizing enterocolitis]. / Aproximacní strevní anastomóza pri multifokální nekrotizující enterokolitide u novorozence.

Multifocal necrotizing enterocolitis (NEC) may result in extensive bowel necrosis and short bowel syndrome. Authors present case report of premature newborn (BW 1700 g, gestational age 30 w.) where an extensive multisegmental NEC of small and large intestine was found during first explorative laparotomy. Proximal jejunostomy 28 cm beyond ligament of Treitz was performed and the rest of involved intestine was left in situ. After 48 hours multiple small bowel resections were performed leaving 12 cm of small intestine (5 short segments) distal to the jejunostomy. Five approximative anastomoses were performed to restore continuity among these segments and ileocaecal valve. Each of approximative anastomosis was constructed with limited number of 4-6 interrupted stitches and all anastomoses healed without complication. Intestinal continuity between proximal jejunostomy and the reconstructed segment of ileum was reestablished nine weeks later. Total length of small bowel was 50 cm. The patient was discharged at the age of 5 months weighing 4145 g with supplemental pareneteral nutrition. The technique of rapid approximative anastomosis may contribute to save maximal intestinal length in cases with the risk of short bowel syndrome.

"Daisy-like" crystals: A rare and unknown type of urinary crystal.

BACKGROUND: Crystals are well known structures of urinary sediment, most of which are identified by the combined knowledge of crystal morphology, birefringence features at polarized light, and urine pH. In this paper, we report on a cohort of subjects whose urine contained a very rare type of crystal, which we first described in 2004 and which, based on its peculiar morphology, we define as "daisy-like crystal" (DLcr). METHODS: Reports on DLcr were spontaneously sent to our laboratory over a 10.5-year period by different laboratory professionals and by one veterinary clinician who, in their everyday work, had come across DLcr. After the examination of DLcr images submitted, a number of other information were requested and partly obtained. RESULTS: DLcr were found in 9 human beings in 7 different laboratories, located in 4 countries (Italy, Belgium, Croatia, France). DLcr were found mostly in female (8/9), at all ages (3.5 to 93years), mostly in alkaline urine (pH6.0 to 7.5), at variable specific gravity values (1.010 to 1.030), either as isolated particles (2/8) or in association with other crystals (5/8) and/or leucocytes or bacteria (3/8). In addition, DLcr were found in the urine of a 1-year-old dog, examined in a veterinary clinic of Czech Republic. In 3 cases, DLcr were identified by manual microscopy, while in 7 cases by automated urine sediment analyzers. CONCLUSIONS: This paper confirms the possible presence in the urine of DLcr. However, further cases are needed to clarify their frequency, clinical meaning, and composition.

Why adult mammalian intrafusal and extrafusal fibers contain different myosin heavy-chain isoforms.

Multiple isoforms of the contractile protein myosin are present in mammalian skeletal muscles. The diversity of the heavy-chain subunits of myosin (MyHCs) in intrafusal fibers is thought to reflect a pathway of differentiation that is unique to muscle spindles. In fact, intrafusal MyHCs are developmental isoforms expressed by the prenatal precursors of both intrafusal and extrafusal fibers. In adult limbs, developmental MyHCs persist in intrafusal, but not extrafusal fibers principally due to the afferent neurons that arrest their maturational replacement by MyHCs associated with faster shortening velocities. The slow shortening velocities that are characteristic of developmental MyHCs might be adaptive for precise calibration of muscle spindles as sense organs.

Mechanistic insights into very slow conduction in branching cardiac tissue: a model study.

It is known that branching strands of cardiac tissue can form a substrate for very slow conduction. The branches slow conduction by acting as current loads drawing depolarizing current from the main strand ("pull" effect). It has been suggested that, upon depolarization of the branches, they become current sources reinjecting current back into the strand, thus enhancing propagation safety ("push" effect). It was the aim of this study to verify this hypothesis and to assess the contribution of the push effect to propagation velocity and safety. Conduction was investigated in strands of Luo-Rudy dynamic model cells that branch from either a single branch point or from multiple successive branch points. In single-branching strands, blocking the push effect by not allowing current to flow retrogradely from the branches into the strand did not significantly increase the branching-induced local propagation delay. However, in multiple branching strands, blocking the push effect resulted in a significant slowing of overall conduction velocity or even in conduction failure. Furthermore, for certain slow velocities, the safety factor for propagation was higher when slow conduction was caused by branching tissue geometry than by reduced excitability without branching. Therefore, these results confirm the proposed "pull and push" mechanism of slow, but nevertheless robust, conduction in branching structures. Slow conduction based on this mechanism could occur in the atrioventricular node, where multiple branching is structurally present. It could also support reentrant excitation in diseased myocardium where the substrate is structurally complex.

Power-law behavior of beat-rate variability in monolayer cultures of neonatal rat ventricular myocytes.

It is known that extracardiac factors (nervous, humoral, and hemodynamic) participate in the power-law behavior of heart-rate variability. To assess whether intrinsic properties of cardiac tissue might also be involved, beat-rate variability was studied in spontaneously beating cell cultures devoid of extracardiac influences. Extracellular electrograms were recorded from monolayer cultures of neonatal rat ventricular myocytes under stable incubating conditions for up to 9 hours. The beat-rate time series of these recordings were examined in terms of their Fourier spectra and their Hurst scaling exponents. A non-0 Hurst exponent was found in 21 of 22 preparations (0.29+/-0.09; range, 0.11 to 0.45), indicating the presence of fractal self-similarity in the beat-rate time series. The same preparations exhibited power-law behavior of the power spectra with a power-law exponent of -1.36+/-0.24 (range, -1.04 to -1.96) in the frequency range of 0.001 to 1 Hz. Furthermore, it was found that the power-law exponent was nonstationary over time. These results indicate that the power-law behavior of heart-rate variability is determined not only by extracardiac influences but also by components intrinsic to cardiac tissue. Furthermore, the presence of power-law behavior in monolayer cultures of cardiomyocytes suggests that beat-rate variability might be determined by the complex nonlinear dynamics of processes occurring at the level of the cellular network, eg, interactions among a large number of cell oscillators or metabolic regulatory systems.

Optical recording of impulse propagation in designer cultures. Cardiac tissue architectures inducing ultra-slow conduction.

It has long been established that slow conduction constitutes one of the key mechanisms in the generation of cardiac arrhythmias. Also, it has been recognized that alterations in the cellular architecture of cardiac tissue can contribute to slow conduction. Based on the recent development of an experimental system permitting both the design of geometrically defined cardiac tissue structures in culture and the measurement of impulse propagation at the cellular level, we investigated the extent of conduction slowing along a tissue structure consisting of a cell strand releasing multiple side branches. This structure, which can functionally be looked upon as a series of interconnected current-to-load mismatches, gave rise to ultra-slow conduction (1-2 cm/s) that displayed a high margin of safety due to a "pull" and "push" effect exerted by the side branches on electrotonic current flow along the main strand. Under physiological conditions, such branching structures might contribute to slow conduction in the AV-node and, under pathophysiological conditions, to the precipitation of reentrant arrhythmias within minuscule tissue regions in a structurally remodeled myocardium. The results illustrate that the combination of patterned growth techniques and optical recording of transmembrane voltage are ideally suited to characterize systematically the relationship between tissue structure and impulse conduction.