Mechanisms of magnetic stimulation of central nervous system neurons

Transcranial magnetic stimulation (TMS) is a stimulation method in which a magnetic coil generates a magnetic field in an area of interest in the brain. This magnetic field induces an electric field that modulates neuronal activity. The spatial distribution of the induced electric field is determined by the geometry and location of the coil relative to the brain. Although TMS has been used for several decades, the biophysical basis underlying the stimulation of neurons in the central nervous system (CNS) is still unknown. To address this problem we developed a numerical scheme enabling us to combine realistic magnetic stimulation (MS) with compartmental modeling of neurons with arbitrary morphology. The induced electric field for each location in space was combined with standard compartmental modeling software to calculate the membrane current generated by the electromagnetic field for each segment of the neuron. In agreement with previous studies, the simulations suggested that peripheral axons were excited by the spatial gradients of the induced electric field. In both peripheral and central neurons, MS amplitude required for action potential generation was inversely proportional to the square of the diameter of the stimulated compartment. Due to the importance of the fiber's diameter, magnetic stimulation of CNS neurons depolarized the soma followed by initiation of an action potential in the initial segment of the axon. Passive dendrites affect this process primarily as current sinks, not sources. The simulations predict that neurons with low current threshold are more susceptible to magnetic stimulation. Moreover, they suggest that MS does not directly trigger dendritic regenerative mechanisms. These insights into the mechanism of MS may be relevant for the design of multi-intensity TMS protocols, may facilitate the construction of magnetic stimulators, and may aid the interpretation of results of TMS of the CNS.     

Biometry based ageing of nestling Indian Spotted Owlets ( Athene brama brama)

Biometric analysis helps in sex differentiation, understanding development and for studies of avian biology such as foraging ecology, evolutionary ecology, and survivorship. We suggest that biometry can also be a reliable, practical and inexpensive tool to determine the age of nestlings in the field by non-invasive methods. As an example we studied the biometry of wing, culmen, talon, tarsus and body mass of nestling southern Indian Spotted Owlets (Athene brama brama). Based on the growth pattern analysis using logistic growth model, discriminant analysis and CHAID (Chi-squared Automatic Interaction Detection) based decision tree, we show that biometry of nestling Spotted Owlets is an easy, reliable and inexpensive method to determine nestling age and to assess growth rate and relative nutritional status. These biometric parameters also allow us to predict their ability to initiate first flight from the nest site. This method is described here for the first time and we postulate that such charts can be devised for other avian species as well, so as to assist conservation biologists and bird rescuers.     

Following cell-fate in E. coli after infection by phage lambda

The system comprising bacteriophage (phage) lambda and the bacterium E. coli has long served as a paradigm for cell-fate determination. Following the simultaneous infection of the cell by a number of phages, one of two pathways is chosen: lytic (virulent) or lysogenic (dormant). We recently developed a method for fluorescently labeling individual phages, and were able to examine the post-infection decision in real-time under the microscope, at the level of individual phages and cells. Here, we describe the full procedure for performing the infection experiments described in our earlier work. This includes the creation of fluorescent phages, infection of the cells, imaging under the microscope and data analysis. The fluorescent phage is a "hybrid", co-expressing wild- type and YFP-fusion versions of the capsid gpD protein. A crude phage lysate is first obtained by inducing a lysogen of the gpD-EYFP (Enhanced Yellow Fluorescent Protein) phage, harboring a plasmid expressing wild type gpD. A series of purification steps are then performed, followed by DAPI-labeling and imaging under the microscope. This is done in order to verify the uniformity, DNA packaging efficiency, fluorescence signal and structural stability of the phage stock. The initial adsorption of phages to bacteria is performed on ice, then followed by a short incubation at 35°C to trigger viral DNA injection. The phage/bacteria mixture is then moved to the surface of a thin nutrient agar slab, covered with a coverslip and imaged under an epifluorescence microscope. The post-infection process is followed for 4 hr, at 10 min interval. Multiple stage positions are tracked such that ~100 cell infections can be traced in a single experiment. At each position and time point, images are acquired in the phase-contrast and red and green fluorescent channels. The phase-contrast image is used later for automated cell recognition while the fluorescent channels are used to characterize the infection outcome: production of new fluorescent phages (green) followed by cell lysis, or expression of lysogeny factors (red) followed by resumed cell growth and division. The acquired time-lapse movies are processed using a combination of manual and automated methods. Data analysis results in the identification of infection parameters for each infection event (e.g. number and positions of infecting phages) as well as infection outcome (lysis/lysogeny). Additional parameters can be extracted if desired.     

Role of c-Abl in the DNA damage stress response

c-Abl has been implicated in many cellular processes including differentiation, division, adhesion, death, and stress response, c-Abl is a latent tyrosine kinase that becomes activated in response to numerous extra- and intra-cellular stimuli. Here we briefly review the current knowledge about c-Abl involvement in the DNA-damage stress response and its implication on cell physiology.     

20S proteasomal degradation of ornithine decarboxylase is regulated by NQO1

Ornithine decarboxylase (ODC), a key enzyme in the biosynthesis of polyamines, is a very labile protein. ODC is a homodimeric enzyme that undergoes ubiquitin-independent proteasomal degradation via direct interaction with antizyme, a polyamine-induced protein. Binding of antizyme promotes the dissociation of ODC homodimers and marks ODC for degradation by the 26S proteasomes. We describe here an alternative pathway for ODC degradation that is regulated by NAD(P)H quinone oxidoreductase 1 (NQO1). We show that NQO1 binds and stabilizes ODC. Dicoumarol, an inhibitor of NQO1, dissociates ODC-NQO1 interaction and enhances ubiquitin-independent ODC proteasomal degradation. We further show that dicoumarol sensitizes ODC monomers to proteasomal degradation in an antizyme-independent manner. This process of NQO1-regulated ODC degradation was recapitulated in vitro by using purified 20S proteasomes. Finally, we show that the regulation of ODC stability by NQO1 is especially prominent under oxidative stress. Our findings assign to NQO1 a role in regulating ubiquitin-independent degradation of ODC by the 20S proteasomes.     

Are the low protein requirements of nectarivorous birds the consequence of their sugary and watery diet? A test with an omnivore

Nectar-feeding birds have remarkably low nitrogen requirements. These may be due either to adaptation to a low-protein diet or simply to feeding on a fluid diet that minimizes metabolic fecal nitrogen losses. We measured minimal nitrogen requirements (MNR) and total endogenous nitrogen loss (TENL) in the omnivorous European starling Sturnus vulgaris, fed on an artificial nectar-like fluid diet of varying concentrations of sugar and protein. The MNR and TENL of the birds were similar and even slightly higher than allometrically expected values for birds of the starlings' mass (140% and 103%, respectively). This suggests that the low measured nitrogen requirements of nectar-feeding birds are not simply the result of their sugary and watery diets but a physiological adaptation to the low nitrogen input. We also measured the effect of water and protein intake on the nitrogenous waste form in the excreta and ureteral urine in European starlings. Neither high water intake nor low protein intake increased the fraction of nitrogen excreted as ammonia. Ammonia was excreted at consistently low levels by the starlings, and its concentration was significantly higher in ureteral urine than in excreta. We hypothesize that ureteral ammonia was reabsorbed in the lower intestine, indicating a postrenal modification of the urine.