Pulmonary vein firing initiating atrial fibrillation in the horse: Oversized dimensions but similar mechanisms

Atrial fibrillation is triggered by the pulmonary veins in humans. Although atrial fibrillation is known to occur in other species, the mechanisms of disease in these are not known. Here we present evidence for pulmonary vein triggers in the horse, where 3D HD Grid mapping was undertaken in the conscious state in the absence of fluoroscopy.     

Afternystagmus in darkness after suppression of optokinetic nystagmus: an interaction of motion aftereffect and retinal afterimages

The afternystagmus that occurs in the dark after gaze fixation during optokinetic stimulation is directed in the opposite direction relative to the previous optokinetic stimulus. The mechanism responsible for such afternystagmus after suppression of optokinetic nystagmus (ASOKN) is unclear. Several hypotheses have been put forward to explain it, but none is conclusive. We hypothesized that ASOKN is driven by the interaction of two mechanisms: (1) motion-aftereffect (MAE)-induced eye movements and (2) retinal afterimages (RAIs) produced by fixation during the suppression of optokinetic nystagmus (OKN). We examined the correlation among ASOKN, MAE-induced eye movements, and RAIs in healthy subjects. Adapting stimuli consisted of moving random dot patterns and a fixation spot and their brightness was adjusted to induce different RAI durations. Test patterns were a stationary random dot pattern (to test for the presence of a MAE), a dim homogeneous background (to test for MAE driven eye movements), and a black background (to test for ASOKN and RAIs). MAEs were reported by 16 out of 17 subjects, but only 7 out of 17 subjects demonstrated MAE-induced eye movements. Importantly, ASOKN was only found when these seven subjects reported a RAI after suppression of OKN. Moreover, the duration of ASOKN was longer for high-brightness stimuli compared with low-brightness stimuli, just as RAIs persist longer with increasing brightness. We conclude that ASOKN results from the interaction of MAE-induced eye movements and RAIs.     

Bile acids increase the activity of the epithelial Na+ channel

The epithelial Na⁺ channel (ENaC) is a key regulator of Na⁺ absorption in various epithelia including the distal nephron and the distal colon. ENaC is a constitutively active channel, but its activity is modulated by a number of mechanisms. These include proteolytic activation, ubiquitination and cell surface expression, phosphorylation, intracellular Na⁺ concentration, and shear stress. ENaC is related to the bile acid-sensitive ion channel (BASIC), a channel that is expressed in the epithelial cells of bile ducts. BASIC is activated by millimolar concentrations of extracellular bile acids. Bile acids are synthesized by the liver and secreted into the duodenum to aid lipolysis. A large fraction of the secreted bile acids is absorbed by the ileum and recirculated into the liver, but a small fraction passes the colon and is excreted. Bile acids can influence the ion transport processes in the intestinal tract including the colon. In this study, we show that various bile acids present in rat bile potently and reversibly increase the activity of rat ENaC expressed in Xenopus oocytes, suggesting that bile acids are natural modulators of ENaC activity.     

In vitro and in vivo Biocompatibility of Alginate Dialdehyde/Gelatin Hydrogels with and without Nanoscaled Bioactive Glass for Bone Tissue Engineering Applications

In addition to good mechanical properties needed for three-dimensional tissue engineering, the combination of alginate dialdehyde, gelatin and nano-scaled bioactive glass (45S5) is supposed to combine excellent cellular adhesion, proliferation and differentiation properties, good biocompatibility and predictable degradation rates. The goal of this study was to evaluate thein vitro and in vivo biocompatibility as a first step on the way to its use as a scaffold in bone tissue engineering. In vitro evaluation showed good cell adherence and proliferation of bone marrow derived mesenchymal stem cells seeded on covalently crosslinked alginate dialdehyde-gelatin (ADA-GEL) hydrogel films with and without 0.1% nano-Bioglass^®(nBG). Lactate dehydrogenase (LDH)- and mitochondrial activity significantly increased in both ADA-GEL and ADA-GEL-nBG groups compared to alginate. However, addition of 0.1% nBG seemed to have slight cytotoxic effect compared to ADA-GEL. In vivo implantation did not produce a significant inflammatory reaction, and ongoing degradation could be seen after four weeks. Ongoing vascularization was detected after four weeks. The good biocompatibility encourages future studies using ADA-GEL and nBG for bone tissue engineering application.     

Overtone-based pitch selection in hermit thrush song: Unexpected convergence with scale construction in human music

Many human musical scales, including the diatonic major scale prevalent in Western music, are built partially or entirely from intervals (ratios between adjacent frequencies) corresponding to small-integer proportions drawn from the harmonic series. Scientists have long debated the extent to which principles of scale generation in human music are biologically or culturally determined. Data from animal “song” may provide new insights into this discussion. Here, by examining pitch relationships using both a simple linear regression model and a Bayesian generative model, we show that most songs of the hermit thrush (Catharus guttatus) favor simple frequency ratios derived from the harmonic (or overtone) series. Furthermore, we show that this frequency selection results not from physical constraints governing peripheral production mechanisms but from active selection at a central level. These data provide the most rigorous empirical evidence to date of a bird song that makes use of the same mathematical principles that underlie Western and many non-Western musical scales, demonstrating surprising convergence between human and animal “song cultures.” Although there is no evidence that the songs of most bird species follow the overtone series, our findings add to a small but growing body of research showing that a preference for small-integer frequency ratios is not unique to humans. These findings thus have important implications for current debates about the origins of human musical systems and may call for a reevaluation of existing theories of musical consonance based on specific human vocal characteristics.Many human musical scales, including the diatonic major scale prevalent in Western music, are built partially or entirely from intervals (ratios between adjacent frequencies) corresponding to small-integer ratios drawn from the harmonic series (1). A long-running debate concerns the extent to which principles underlying the structure of human musical scales derive from biological aspects of auditory perception and/or vocal production or are historical cultural “accidents” (2–4). The songs of nonhuman animals, such as birds or whales, potentially offer a valuable perspective on this debate. On the one hand, features of human music that are culturally bound, or dependent on specific characteristics of the human voice or auditory system, should be absent in animal vocalizations. On the other hand, aspects of human music observed in the vocalizations of other species seem likely to be partially determined by general physical or biological constraints rather than solely by cultural practices. Such shared features would complement recent research suggesting that common motor constraints shape both human song and that of some bird species (5).The physical principles underlying vocal production in songbirds are well understood (6–10) and do not differ fundamentally from those of other vertebrates. Sound is produced by tissue vibrations in the syrinx, a bird-specific organ located at the base of the trachea. Flow-driven vibrations of fleshy membranes within the syrinx (in songbirds, the medial and lateral labia) generate a periodic source signal that is filtered by the air column within the trachea and mouth and then emitted to the environment. These principles are important in formulating various alternative hypotheses considered below.Naturalists have long wondered whether birdsong could be said to have musical properties (11–13). However, early studies on pitch selection tended to be anecdotal, based on a small sample size, or lacking in analytical rigor. Two more recent studies specifically comparing pitch selection in bird song and human musical scales concluded that birdsong does not make preferential use of musical intervals found in commonly used Western musical scales (14, 15). However, because these studies each only examined one species [the white-throated sparrow (Zonotrichia albicollis) and the nightingale wren (Microcerculus philomela), respectively], a conclusion that birdsong in general does not exhibit musical properties seems premature. Indeed, other studies have shown preferential use of consonant intervals in tropical boubou shrikes (Laniarius aethiopicus) (16) and musician wrens (Cyphorhinus arada) (17), although in the first case no rigorous statistical analysis was presented.Here, we investigated songs of the hermit thrush (Catharus guttatus), a medium-sized North American songbird whose famously “musical”-sounding song has attracted the attention of ornithologists and musicians alike (18) but has not yet been subjected to detailed pitch analysis. Its songs are composed of elements (the smallest unit of song construction, seen as continuous uninterrupted traces on spectrograms) that may exhibit either a variable pitch, such as trills and slides, or a stable pitch—pure, non-frequency-modulated, “flutelike” sounds. These stable sounds, which we refer to as “notes” (Fig. 1), are characterized by strong fundamental frequencies and very weak higher harmonics, making them ideally suited for an analysis of pitch relationships (15). Males typically sing 6–10 different song types, defined as nearly identical sequences of elements, durations, and frequencies. In a number of early- and mid-20th-century studies, hermit thrush song was variously attributed with use of major, minor, and pentatonic scales (19, 20) and claimed to follow the overtone series (21). However, these early studies again suffered from small sample sizes and anecdotal reporting and were not based on rigorous acoustic analysis. More recent hermit thrush studies have focused on regional differences and song-type ordering, rather than pitch selection (22, 23).Open in a separate windowFig. 1.Song of the hermit thrush (C. guttatus). One song type of a single male hermit thrush, illustrating the various elements that can be observed in songs of this species. Only “notes” (elements with stable pitch) were analyzed in this study because the other element types have no clearly defined or measurable pitch.Here we tested the overtone hypothesis, which predicts that the frequencies of the individual song notes are integer multiples (harmonics) of an implied (but not actually sung) base frequency (hereafter f_i). This hypothesis seems plausible because, unlike some previous claims, it does not attribute human-specific music-theoretical concepts to hermit thrush song. Moreover, the subjective impression of trained musicians listening to hermit thrush songs (played at one-sixth of the original speed to shift the speed and frequency of the songs into a range more suitable for human hearing) was that most notes indeed seemed to follow an overtone series (see Fig. 2 and Audio File S1 for the corresponding sound example). However, determining whether a set of notes are harmonics of a frequency not present in the set requires a rigorous procedure to estimate and evaluate f_i. To this end, we used two different statistical approaches, an ordinary least-squares regression model and a generative Bayesian estimator. Both approaches were used to test the hypothesis that a song is an exchangeable sequence of frequencies that are integer multiples of some implied f_i, versus the null hypothesis that songs are generated by drawing frequencies out of a random log-normal distribution (see Materials and Methods for details). By using a Bayesian approach in addition to the least-squares regression model we evaluate whether our analyses represent a rigorous test of our overtone hypothesis and not simply a post hoc explanation that minimizes an error measure by “memorizing” the data. These properties make the Bayesian evaluation statistically more rigorous than least-squares fitting.Open in a separate windowFig. 2.Frequency distribution of a hermit thrush song compared with an overtone series. (A) Notes of a hermit thrush song. (B) The same notes rearranged in ascending order to show how they correspond to overtones 3, 4, 5, and 6 of an overtone series fitted to the frequencies corresponding to these notes (the complete stacked overtone series is shown on the right).     

Genetic activation of pyruvate dehydrogenase alters oxidative substrate selection to induce skeletal muscle insulin resistance

The pyruvate dehydrogenase complex (PDH) has been hypothesized to link lipid exposure to skeletal muscle insulin resistance through a glucose-fatty acid cycle in which increased fatty acid oxidation increases acetyl-CoA concentrations, thereby inactivating PDH and decreasing glucose oxidation. However, whether fatty acids induce insulin resistance by decreasing PDH flux remains unknown. To genetically examine this hypothesis we assessed relative rates of pyruvate dehydrogenase flux/mitochondrial oxidative flux and insulin-stimulated rates of muscle glucose metabolism in awake mice lacking pyruvate dehydrogenase kinase 2 and 4 [double knockout (DKO)], which results in constitutively activated PDH. Surprisingly, increased glucose oxidation in DKO muscle was accompanied by reduced insulin-stimulated muscle glucose uptake. Preferential myocellular glucose utilization in DKO mice decreased fatty acid oxidation, resulting in increased reesterification of acyl-CoAs into diacylglycerol and triacylglycerol, with subsequent activation of PKC-θ and inhibition of insulin signaling in muscle. In contrast, other putative mediators of muscle insulin resistance, including muscle acylcarnitines, ceramides, reactive oxygen species production, and oxidative stress markers, were not increased. These findings demonstrate that modulation of oxidative substrate selection to increase muscle glucose utilization surprisingly results in muscle insulin resistance, offering genetic evidence against the glucose-fatty acid cycle hypothesis of muscle insulin resistance.Lipid-induced muscle insulin resistance plays a major role in the pathogenesis of type 2 diabetes (T2D), but the cellular mechanisms remain unknown (1, 2). More than 50 y ago Randle et al. (3) postulated the glucose-fatty acid cycle to explain the impairment of insulin-stimulated glucose disposal by fatty acids in muscle. In this model, fat oxidation increases mitochondrial acetyl-CoA/CoA and NADH/NAD⁺ ratios. Acetyl-CoA and NADH allosterically inhibit pyruvate dehydrogenase complex (PDH), the mitochondrial enzyme that links glycolysis to the TCA cycle by converting pyruvate to acetyl-CoA. Additionally, fatty acid-derived acetyl-CoA produces citrate, which inhibits phosphofructokinase. This in turn increases glucose-6-phosphate (G6P), a potent allosteric inhibitor of hexokinase. By these mechanisms, increased fatty acid oxidation was hypothesized to reduce glycolytic flux and prevent further muscle glucose uptake. However, in vivo studies of human skeletal muscle metabolism have challenged the Randle hypothesis. Five hours of a lipid infusion, combined with heparin to activate lipoprotein lipase, raised plasma fatty acids and induced muscle insulin resistance in healthy individuals, yet intramyocellular G6P and glucose concentrations were reduced compared with control glycerol infusion studies, implicating defects in insulin-stimulated glucose transport activity (4, 5). An alternative hypothesis to explain the muscle insulin resistance associated with lipid exposure posits that accumulation of bioactive lipid intermediates initiates signaling cascades that impair insulin action. Lipid species implicated include diacylglycerols (DAGs) (6–10), ceramides (11, 12), and long-chain acyl-CoAs (13). DAG activation of PKC-θ in skeletal muscle has been shown to impair canonical insulin signaling at the level of insulin receptor substrate-1 (IRS-1) tyrosine phosphorylation through increased IRS-1 serine phosphorylation at the 1101 position (2, 6, 7, 14).More recently, incomplete fat oxidation and subsequent accumulation of mitochondrially derived acylcarnitines has been proposed to contribute to lipid-induced muscle insulin resistance (15–17). According to this model, insulin resistance stems from increased fat oxidation, leading to increased conversion of acyl-CoA to medium- and long-chain acylcarnitines, which may mediate insulin resistance via unknown mechanisms. In contrast, short-chain acylcarnitines have been suggested to promote metabolic flexibility. The shortest acylcarnitine, acetylcarnitine, is synthesized from acetyl-CoA and carnitine by carnitine acetyltransferase (CrAT), a mitochondrial matrix enzyme, and is responsible for buffering the mitochondrial acetyl-CoA pool and mitigating acetyl-CoA inhibition of PDH (18). Consistent with the notion that CrAT regulates substrate selection by modulating PDH flux, mice with muscle-specific deletion of CrAT exhibited reduced PDH activity during the fed-to-fasted transition, resulting in glucose intolerance and metabolic inflexibility, a term coined by Kelley and Mandarino (19) to explain the impairment in the ability to adjust fuel oxidation to fuel availability.Although these studies emphasize the importance of PDH in the promotion of metabolic inflexibility, the role of PDH and mitochondrial oxidative substrate selection in the regulation of basal and insulin-stimulated muscle glucose metabolism has not been directly assessed in vivo. To examine this question, we sought to determine whether modulation of oxidative substrate selection in a genetic mouse model with constitutively active PDH activity would affect insulin sensitivity in skeletal muscle.     

Charge separation and charge delocalization identified in long-living states of photoexcited DNA

Base stacking in DNA is related to long-living excited states whose molecular nature is still under debate. To elucidate the molecular background we study well-defined oligonucleotides with natural bases, which allow selective UV excitation of one single base in the strand. IR probing in the picosecond regime enables us to dissect the contribution of different single bases to the excited state. All investigated oligonucleotides show long-living states on the 100-ps time scale, which are not observable in a mixture of single bases. The fraction of these states is well correlated with the stacking probabilities and reaches values up to 0.4. The long-living states show characteristic absorbance bands that can be assigned to charge-transfer states by comparing them to marker bands of radical cation and anion spectra. The charge separation is directed by the redox potential of the involved bases and thus controlled by the sequence. The spatial dimension of this charge separation was investigated in longer oligonucleotides, where bridging sequences separate the excited base from a sensor base with a characteristic marker band. After excitation we observe a bleach of all involved bases. The contribution of the sensor base is observable even if the bridge is composed of several bases. This result can be explained by a charge delocalization along a well-stacked domain in the strand. The presence of charged radicals in DNA strands after light absorption may cause reactions—oxidative or reductive damage—currently not considered in DNA photochemistry.DNA photophysics is crucial for the understanding of light-induced damage of the genetic code (1). The excited state of single DNA bases is known to decay extremely fast on the subpicosecond time scale, predominantly via internal conversion (2, 3). This ultrafast decay is assumed to suppress destructive decay channels, thereby protecting the DNA from photodamage and avoiding disintegration of the genetic information. In contrast to this ultrafast deactivation of single nucleobases, the biological relevant DNA strands show further long-living states (4, 5). Several explanations for these long-living states and the size of their spatial extent have been discussed in the literature (5–9). Delocalized excitons (9); excitons that decay to charge-separated states or neutral excimer states (10, 11); exciplexes located on two neighboring bases (5, 8, 12, 13); or even excited single bases, where steric interactions in the DNA strand impedes the ultrafast decay (14), have been proposed. Further computations suggest a decay of an initially populated delocalized exciton to localized neutral or charged excimer states (15–17). However, to our knowledge, a final understanding of the nature of these long-living states has not been reached. Related experiments were performed in the last decade to investigate charge transport processes in DNA, motivated by DNA electronics and oxidative damage (18, 19). Charge transport was initiated by photoexcitation of modified DNA bases or chromophores and followed by transient absorption (20–23). The transport mechanism was described by charge-hopping, superexchange, or transfer of charge along delocalized domains in DNA (18).Until now, most experimental investigations of the long-living state were performed with transient absorption spectroscopy in the UV-visible (UV/Vis) regime (5, 9, 12) or with time-resolved fluorescence (10, 24, 25). Due to the broad, featureless, and overlapping absorption bands of the different DNA bases in this spectral region, it is difficult to investigate the molecular origin of the long-living states using these methods. A further drawback is the unselective and simultaneous excitation of several bases used in most experiments. To circumvent these problems, we used for the present study well-defined oligonucleotides, which enable selective excitation of one single base. Observation of the long-living excited states was performed via time-resolved IR spectroscopy, which can profit from the many “fingerprint” vibrational bands (26, 27). IR spectroscopy is able to distinguish between different DNA bases and their molecular states. It can also reveal changes in the electronic structure and identify charge-separated states.In this study we used single-stranded DNA, in which π stacking between neighboring bases leads to structured domains, similar to the structure in a double helix (28). This interaction is known to be crucial for the long-living states (5). The investigation of single-stranded DNA enables us to construct special sequences, where only one base can be selectively excited. We used the natural bases 2′-deoxyuridine (U), 2′-deoxyadenosine (A), 5-methyl-2′-deoxycytidine (mC), and 2′-deoxyguanosine (G). The nucleobase U occurs naturally in RNA and is similar to the DNA base thymine but shows a blue-shifted absorbance spectrum. mC occurs with a frequency of 4–5% in mammalian DNA (29) and plays an important role as an epigenetic marker (30). The UV/Vis absorbance of mC and G are red-shifted in comparison with A and U, which allows selective excitation at 295 nm in oligonucleotides consisting of mC, A, and U (Fig. 1 A and B) or G and A. This selectivity can only be obtained in single-stranded DNA because G and its complementary base mC have overlapping absorbance bands in the UV range (Fig. S1). Selectivity in probing is based on the significant differences in the IR-absorption spectra of these bases, which display distinct marker bands for each base (Fig. 1 A and E).Open in a separate windowFig. 1.Selective excitation of mC in mCUA and probing of characteristic A, U, and mC marker bands in the IR. (A, B, and E) Picosecond UV light pulses at 295 nm allow selective excitation of mC (shown in bold) in mixed DNA sequences consisting of mC, U, and A. (B) Absorbance spectra of 2′-deoxyadenosine monophosphate (A), 2′-deoxy-5-methylcytidine (mC), and uridine monophosphate (U). (C) Time-resolved absorption difference (color-coded) plotted vs. wavenumber and delay time for mCUA and (D) for a mixture of the corresponding monomers. (E) Probing the individual contribution of each base is possible in the IR at 1,625 cm⁻¹ (A), 1,655 cm⁻¹ (U), and 1,667 cm⁻¹ (mC) (marked by dashed lines). (F) Transients at 1,667 cm⁻¹ for mCUA and the mixture of monomers.With the combination of selective excitation and selective probing we are able to elucidate the nature of the long-living states in DNA strands. Investigation of dinucleotides clearly shows that light absorption in DNA leads to charge separation between stacked neighboring bases, which recombine on the 100-ps time scale. In longer oligonucleotides we observe simultaneous bleach of several bases, which points to a delocalization of the charges along the strand. Our results show that charge transfer in DNA is a natural process, induced by UV-light absorption of DNA.