Cortical Modulation of the Transient Visual Response at Thalamic Level: A TMS Study

The transient visual response of feline dorsal lateral geniculate nucleus (dLGN) cells was studied under control conditions and during the application of repetitive transcranial magnetic stimulation at 1 Hz (rTMS@1Hz) on the primary visual cortex (V1). The results show that rTMS@1Hz modulates the firing mode of Y cells, inducing an increase in burst spikes and a decrease in tonic firing. On the other hand, rTMS@1Hz modifies the spatiotemporal characteristics of receptive fields of X cells, inducing a delay and a decrease of the peak response, and a change of the surround/center amplitude ratio of RF profiles. These results indicate that V1 controls the activity of the visual thalamus in a different way in the X and Y pathways, and that this feedback control is consistent with functional roles associated with each cell type

Regulation of cyclic GMP metabolism in toad photoreceptors. Definition of the metabolic events subserving photoexcited and attenuated states

Photoreceptor metabolism of cGMP and its regulation were characterized in isolated toad retinas by determining the intensity and time dependence of light-induced changes in the following metabolic parameters: cGMP hydrolytic flux determined by the rate of 18O incorporation from 18O-water into retinal guanine nucleotide α-phosphoryls; changes in the total (protein-bound and unbound) concentrations of the guanine nucletodie metabolic intermediates; and changes in the concentration of metabolic (unbound) GDP calculated from the fraction of the α-GDP that undergoes labeling with 18O. The latter is interpreted to reflect the state of the equilibrium between GDP-GTP-complexed forms of G-protein. With narrow band 500 nm light that preferentially stimulates ared rod photoreceptors, a range of intensities covering approximately 5 log units produced increases of over 10-fold in cGMP metabolic flux. However, the characteristics of the cGMP metabolic response over the first 2.5 log units of intensity are readily distinguishable from those at higher intensities which exhibit progressive attenuation by an intensity- and time-dependent process. Over the range of low intensities (0.6-3 log photons·μm-2·s-1) the metabolic response is characterized by 1) increases in cGMP hydrolytic flux of up to 8-fold as a logarithmic function of intensity of photic stimulation that are sustained for at least 200 s; 2) small increases or no change in the concentration of total cGMP; 3) large increases of up to 10-fold in the concentration of metabolically active GDP as a linear function of intensity with no significant change in the tissue concentrations of total GDP or GTP; and 4) amplification of the photosignal by the metabolism of approximately 10,000 molecules of cGMP per photoisomerization with the major site of amplification at the level of the interaction of bleached rhodopsin with G-protein. The attenuated cGMP hydrolytic response observed in a high light intensity range (3-5.39 log photons·μm-2·s-1) exhibits the following characteristics: 1) a progressive decline in the magnitude of the light-accelerated cGMP hydrolytic flux response with respect to the photon flux which at the highest intensities leads to relative declines in the cGMP flux after 20 s of illumination and more pronounced decreases after 200 s of illumination; 2) progressive decreases of as much as 25% at 20 s and up to 50% at 200 s in the total cGMP concentration that are proportional to the suppression observed in light-enhanced cGMP hydrolytic flux; and 3) decreases from the maximal levels achieved in the concentration of metabolic (unbound) GDP with no change in total GDP or GTP concentration. The widely held view of a light-activated cascade directed only toward phosphodiesterase cannot account for any of the above features of cGMP metabolism in the intact photoreceptor. To sustain a cGMP flux that is severalfold greater than that in the dark-adapted state, continuous illumination must activate the entire cGMP metabolic cycle, which consists of phosphodiesterase, guanylyl cyclase, and the enzymes phosphorylating GMP and GDP. This mechanism is also consistent with the observation that concentrations of cGMP, GDP, and GTP remain nearly constant during low intensity photic stimulation. To produce the time-dependent attenuation in light-activated flux with the accompanying reductions in total cGMP concentration, intense and extended illumination must result in a gradual down-regulation of light-activated guanylyl cyclase. This perspective contradicts the currently held view of the mechanisim, outcome, and purpose of light-induced alterations in cGMP metabolism.link_to_subscribed_fulltex

Adenosine triphosphate utilization rates and metabolic pool sizes in intact cells measured by transfer of 18O from water

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Dynamic Phosphometabolomic Profiling of Human Tissues and Transgenic Models By O-18-Assisted P-31 Nmr and Mass Spectrometry

Next-generation screening of disease-related metabolomic phenotypes requires monitoring of both metabolite levels and turnover rates. Stable isotope O-18-assisted P-31 nuclear magnetic resonance (NMR) and mass spectrometry uniquely allows simultaneous measurement of phosphometabolite levels and turnover rates in tissue and blood samples. The O-18 labeling procedure is based on the incorporation of one O-18 into Pi from [O-18]H2O with each act of ATP hydrolysis and the distribution of O-18-labeled phosphoryls among phosphate-carrying molecules. This enables simultaneous recording of ATP synthesis and utilization, phosphotransfer fluxes through adenylate kinase, creatine kinase, and glycolytic pathways, as well as mitochondrial substrate shuttle, urea and Krebs cycle activity, glycogen turnover, and intracellular energetic communication. Application of expanded O-18-labeling procedures has revealed significant differences in the dynamics of G-6-P[O-18] (glycolysis), G-3-P[O-18] (substrate shuttle), and G-1-P[O-18] (glycogenolysis) between human and rat atrial myocardium. In human atria, the turnover of G-3-P[O-18], which defects are associated with the sudden death syndrome, was significantly higher indicating a greater importance of substrate shuttling to mitochondria. Phosphometabolomic profiling of transgenic hearts deficient in adenylate kinase (AK1-/-), which altered levels and mutations are associated to human diseases, revealed a stress-induced shift in metabolomic profile with increased CrP[O-18] and decreased G-1-P[O-18] metabolic dynamics. The metabolomic profile of creatine kinase M-CK/ScCKmit-/--deficient hearts is characterized by a higher G-6-[O-18]P turnover rate, G-6-P levels, glycolytic capacity, gamma/beta-phosphoryl of GTP[O-18] turnover, as well as beta-[O-18]ATP and beta-[O-18]ADP turnover, indicating altered glycolytic, guanine nucleotide, and adenylate kinase metabolic flux. Thus, O-18-assisted gas chromatography-mass spectrometry and P-31 NMR provide a suitable platform for dynamic phosphometabolomic profiling of the cellular energetic system enabling prediction and diagnosis of metabolic diseases states.Wo

Light-Evoked Responses of the Retinal Pigment Epithelium: Changes Accompanying Photoreceptor Loss in the Mouse

Mutations in genes expressed in the retinal pigment epithelium (RPE) underlie a number of human inherited retinal disorders that manifest with photoreceptor degeneration. Because light-evoked responses of the RPE are generated secondary to rod photoreceptor activity, RPE response reductions observed in human patients or animal models may simply reflect decreased photoreceptor input. The purpose of this study was to define how the electrophysiological characteristics of the RPE change when the complement of rod photoreceptors is decreased. To measure RPE function, we used an electroretinogram (dc-ERG)-based technique. We studied a slowly progressive mouse model of photoreceptor degeneration (PrphRd2/+), which was crossed onto a Nyxnob background to eliminate the b-wave and most other postreceptoral ERG components. On this background, PrphRd2/+ mice display characteristic reductions in a-wave amplitude, which parallel those in slow PIII amplitude and the loss of rod photoreceptors. At 2 and 4 mo of age, the amplitude of each dc-ERG component (c-wave, fast oscillation, light peak, and off response) was larger in PrphRd2/+ mice than predicted by rod photoreceptor activity (RmP3) or anatomical analysis. At 4 mo of age, the RPE in PrphRd2/+ mice showed several structural abnormalities including vacuoles and swollen, hypertrophic cells. These data demonstrate that insights into RPE function can be gained despite a loss of photoreceptors and structural changes in RPE cells and, moreover, that RPE function can be evaluated in a broader range of mouse models of human retinal disease

P-31 Nmr Correlation Maps Of O-18/O-16 Chemical Shift Isotopic Effects For Phosphometabolite Labeling Studies

Intramolecular correlations among the O-18-labels of metabolic oligophosphates, mapped by J-decoupled P-31 NMR 2D chemical shift correlation spectroscopy, impart stringent constraints to the O-18-isotope distributions over the whole oligophosphate moiety. The multiple deduced correlations of isotopic labels enable determination of site-specific fractional isotope enrichments and unravel the isotopologue statistics. This approach ensures accurate determination of O-18-labeling rates of phosphometabolites, critical in biochemical energy conversion and metabolic flux transmission. The biological usefulness of the J-decoupled P-31 NMR 2D chemical shift correlation maps was validated on adenosine tri-phosphate fractionally O-18 labeled in perfused mammalian hearts.Wo