Contributors:Iain N. McSherry, Michaela M. Spitaler, Hiromichi Takano, Kim A. Dora
Ca2+ oscillations... Effect of ACh on endothelial cell [Ca2+]i in isolated sheets of cells. (A) Confocal brightfield (top) and fluorescence (bottom) images of an isolated endothelial cell sheet loaded with fluo-4. (B andC) The time course of endothelial cell fluorescence changes in response to 1μM ACh added at the time indicated (by ▴). (B) The average whole-cellular fluorescence intensity over time. The coloured lines correspond to the individual cells marked in (A). Note that the oscillations in fluorescence are partially synchronized. (C) The average multi-cellular relative fluorescence (F/F0) over time. The region of interest for measuring fluorescence intensity was increased to include all the cells depicted in (A). (D) Time course of increases in membrane potential (Em) in response to 3μM ACh added at the time indicated (by ▾). The average resting membrane potential was −16.7±2.4mV (n=3 cells), which was increased to −28.8±3.9mV (n=3 cells) by current injection (to cell 1: black trace; and cell 3: red trace). ACh hyperpolarized these cells to a peak of −67.1±2.1mV (n=3 cells). The response to ACh was reproducible (cell 2: green and blue traces correspond to first and second additions), and in two of the three cells, the membrane potential oscillated (frequency: 0.14±0.02Hz, n=3 responses). (E) The cell marked with the arrow in (A) was used to illustrate the time course of sub-cellular fluorescence changes in response to 1μM ACh (added at ▴). Top panel: Changes in F/F0 in the regions corresponding to the coloured boxes shown over the cell below. Note that the increases in F/F0 were synchronous. Bottom panels: The difference in fluorescence intensity (F−F0) across the length of the cell (white line over image) over time (single oscillation indicated by the bar in the top panel). The dotted line in each panel corresponds to the x-coordinate in the image above (white line), and t=0s. There was no evidence for a Ca2+ wave. Note that the longitudinal axis of the artery was horizontally orientated, as in Fig. 1A. A movie of this experiment is available online (Movie 2, Supplementary material).
... Effect of ACh on endothelial cell [Ca2+]i in pressurized arteries. (A) Confocal fluorescence image of arterial endothelial cells loaded with fluo-4. (B and C) The time course of endothelial cell fluorescence changes in response to 0.3μM ACh added at the time indicated (by ▴). (B) The average whole-cellular fluorescence intensity over time. The coloured lines correspond to the individual cells marked in (A), note that the oscillations in fluorescence were asynchronous. (C) The average multi-cellular relative fluorescence (F/F0) over time. The region of interest for measuring fluorescence intensity was increased to include all the cells in the field of view (>50 cells). Note that the pattern of oscillations was markedly dampened compared to the individual cells, reflecting the asynchronous signals. (D) Summarized data showing the time course of increases in F/F0 in response to 0.3μM ACh (n=10) or 3μM ACh (added at t=0s, n=3). (E) The cell marked with an arrow in (A) was enlarged (image below) and used to illustrate the time course of sub-cellular fluorescence changes in response to 0.3μM ACh (added at ▴). Top panel: changes in F/F0 in the regions corresponding to the coloured boxes shown over the cell below. Note that the increases in F/F0 started moving from right to left, were not uniform, and changed direction. Bottom five panels: The difference in fluorescence intensity (F−F0) across the length of the cell (white line in image) over time (a single oscillation indicated by the bar in the top panel). The dotted line in each panel corresponds to the x-coordinate in the image above (white line), and t=0s. The wave travelled from right to left at a speed of approximately 30μm/s, and appeared to quench at the position of the nucleus. A movie of this experiment is available online (Movie 1, Supplementary material).
... Effect of treatments on the oscillations in endothelial cell [Ca2+]i in pressurized arteries. The effect of the various treatments on the oscillations in endothelial cell [Ca2+]i in response to ACh in pressurized arteries. The values were obtained from the experiments presented in Figs. 1, 3 and 4. The time course following the addition of 0.3μM ACh to the bath was divided into two phases, initial: t=0–30s; and sustained: t=30–100s. The percentage of cells oscillating (A) was calculated from the 16 cells analyzed per artery, and the frequency of oscillations (B) calculated using cells that oscillated in (A).
Contributors:Huikun Wang, Tyler Treadway, Daniel P. Covey, Joseph F. Cheer, Carl R. Lupica
Cocaine-Mobilized 2-AG Inhibits GABA Release and Increases DA Release in NAc via Actions in VTA
(A) Acute cocaine (10 μM) perfusion inhibited GABAB IPSCs and this was partially blocked by AM251 (2 μM). Scale bar, 100 ms, 10 pA.
(B) Cocaine inhibition of GABAB IPSCs was reduced by inhibition of DGL with THL, applied either intracellularly via the pipette (intra-THL) or extracellularly, by addition to the aCSF (extra-THL). Scale bar, 100 ms, 10 pA.
(C) Summary of the experiments shows significant reduction of the cocaine inhibition of GABAB IPSCs by AM251 and THL (n = 5; F(3,36) = 6.35, p = 0.0014, one-way ANOVA; ∗p 0.05, paired t test. Scale bar, 100 ms, 10 pA.
(E) In vivo NAc FSCV current (encoded in color) is plotted against the applied potential (ordinate) and the acquisition time (abscissa). Traces above color plots are extracted currents (normalized to concentration) from the potential where DA is oxidized (∼+0.6 V shown in green), when animals are given intravenous cocaine (1 mg/kg), preceded by an i.c.v. vehicle injection.
(F) Effects of cocaine are reduced in vivo by inhibition of 2-AG synthesis by THL in the VTA (750 ng THL in 500 μl, unilateral and ipsilateral to the recording electrode).
(G) Intra-VTA THL reduces DA transient frequency elicited by i.v. cocaine (F (4,10) = 9.395, p = 0.002, one-way ANOVA; n = 3 rats, ∗∗p frequency without intracranial or systemic injections. In other groups, the intra-VTA injection is indicated first, followed by the i.v. injection condition (e.g., intracranial Veh∖intra-VTA Veh). Note that THL alone (THL/Veh) did not alter DA transient frequency, and THL significantly reduced the effect of i.v. cocaine on DA transients (THL/cocaine, p < 0.001, Bonferroni post hoc test).
See also Figure S1.
... Cocaine-Induced Increase in NAc DA Transients Require CB1R Activation in VTA
(A) Voltammetric current (encoded in color) plotted against the applied carbon-fiber electrode potential (ordinate) and the acquisition time (abscissa). Traces above color plots represent current (normalized to concentration) from the potential where DA is oxidized (∼+0.6 V in green), when animals are given an i.v. bolus of cocaine (1 mg/kg), preceded by intra-VTA vehicle infusion.
(B) Systemic effects of cocaine are attenuated following CB1R blockade in VTA (250 ng rimonabant [Rbt] in 500 μl, unilateral and ipsilateral to the recording electrode).
(C) Effects of all treatments on NAc DA transient frequency (F(4,31) = 29.14, p frequency of DA transients without intracranial or systemic injections. In other groups the intra-VTA injection is indicated first, followed by the i.v. injection (e.g., intracranial Veh∖intra-VTA Veh). Note that Rbt alone (Rbt/Veh) did not alter DA transient frequency, and Rbt significantly reduced the effect of i.v. cocaine on DA transients (Rbt/cocaine, ∗∗∗p < 0.0001, Bonferroni post hoc test).
... Cocaine-Induced Calcium Changes in VTA DA Neurons
(A) Superimposed fluorescent and transmitted light images showing the expression of GCaMP6f in VTA DA neurons 2–3 weeks after injection of the AAV- GCaMP6f construct. Scale bar, 500 μm.
(B) Spontaneous slow calcium oscillations in VTA DA neurons. Left: confocal image of the GCaMP6f signal (200×). Scale bar, 50 μm. Right: example calcium oscillations (ΔF/F0) in the same cell indicated in the dashed square at left. Scale bar, 30 s, 20%.
(C) Sample traces of calcium signals (ΔF/F0) in response to cocaine application (10 μM, indicated in gray). Three types of changes were observed: no change, a decrease in spontaneous oscillations, and the initiation of slow oscillations (SO).
(D) Raster plot of oscillation events in 19 neurons imaged as in (B). Each vertical bar denotes the peak of an oscillation event. Each row exhibits the time course of oscillation change in single neurons.
(E) Cumulative frequencies of spontaneous, cocaine-decreased, and cocaine-initiated slow oscillations (cocaine SO) in calcium signal. The curve shifts left to slower frequencies in neurons whose calcium oscillations were decreased by cocaine (osc decrease versus spontaneous, p = 0.17, K-S test). The frequency of cocaine-initiated SO was significantly lower than that of spontaneous oscillations (cocaine-initiated versus spontaneous, p oscillation, n = 43 neurons; oscillation decrease, n = 19 neurons; cocaine SO, n = 29 neurons recorded in eight brain slices from six rats.
(F) Cumulative distribution of half-widths of spontaneous, cocaine-decreased, and cocaine-initiated SO. Cocaine-initiated SO exhibit longer half-widths than spontaneous oscillations. (cocaine SO versus spontaneous, p oscillation, n = 43 neurons; oscillation decrease, n = 19 neurons; cocaine SO, n = 29 neurons recorded in eight brain slices from six rats.
(G) Increased correlation of calcium responses among neurons (same 19 neurons as D) during cocaine perfusion. Cross correlations among all neuron pairs were calculated, and the peak correlation coefficient (r = 0 to 1) is color-coded to form the heat map. The data indicate that VTA DA neuron calcium oscillations are more highly synchronized by cocaine.
(H) Cocaine-induced calcium oscillations depend on mGluR1, α1-adrenergic receptors, and internal calcium stores. The frequency of spontaneous calcium oscillations is reduced by JNJ16259685 (500 nM), but is not altered by HEAT (1 μM), U73122, or thapsigargin (thaps, 2 μM), respectively (F(4,265) = 3.806, p = 0.005, one-way ANOVA; control versus JNJ: p frequency of spontaneous Ca2+ oscillation from one neuron; the box and vertical lines indicate quartiles, minimal, and maximal frequencies. The percentage of neurons exhibiting spontaneous oscillations is shown for each treatment group.
(I) Cumulative distributions of the frequencies of spontaneous oscillations are only affected by JNJ (p = 0.0037, Kolmogorov–Smirnov test).
(J) Summary of the percentage of DA neurons responding to cocaine under control, JNJ16259685, HEAT, U73122, and thapsigargin treatments. Numbers denote the percentage of neurons demonstrating indicated changes upon cocaine treatment. All treatments decreased the cocaine-induced Ca2+ oscillation.
(K) Pre-treatment of slices with thapsigargin (2 μM) significantly attenuates the inhibition of GABAB IPSCs by cocaine (n = 7; ∗p < 0.05, unpaired t test). Inset at right shows averaged electrically evoked GABAB IPSC traces (n = 5) under control conditions (top) and after treatment with thapsigargin. Control sweeps are shown in black; those obtained during cocaine application are shown in gray. Scale bar, 100 ms, 10 pA.
See also Movie S1.
Contributors:Gonzalo J. Revuelta, Subramaniam Uthayathas, Amy E. Wahlquist, Stewart A. Factor, Stella M. Papa
NHP FOG developed in 14 out of 29 monkeys (48%) with stable parkinsonism, as follows: 2 monkeys were from the prospective examination group of 6 animals, and 12 from the retrospective review group of 23 animals (videotapes of NHP FOG were available on 5 cases in retrospective review). NHP FOG were sudden episodes lasting from several seconds to approximately 1min, with clear evidence that the monkeys were able to walk without freezing before and after each episode (see videotapes in Supplementary materials). The frequency of NHP FOG was not documented, however, it was consistently observed in successive morning “off” state evaluations. We could not identify any specific provoking factors, although typical provoking factors for humans were not utilized due to the inherent difficulties of evaluating caged monkeys. In some cases there was evidence of hesitation or trembling of the hind limb. In addition, when the monkey's gait froze, there appeared to be an associated generalized akinesia. NHP FOG could be observed during walking and climbing. A clear relation to starting or reaching the destination could not be established because of our inability to determine the animal's intention to stop suddenly or his predetermined destination. Monkeys with FOG also had a tendency to slowly flex the legs until sitting down after they froze, which does not occur in humans. But in the monkey, sitting is an easily reached position from the standing for quadrupedal gait. The individual demographic characteristics and the MPTP treatment in the FOG monkey group (n=14) are presented in detail in Table 1, and those in the non-FOG monkey group (n=15) in Supplementary materials, Table S1.... Analysis of the evolution of FOG episodes in the MPTP-treated monkey. A and B, recordings of leg movements with an accelerometer placed on the back of the leg during the whole duration of FOG episodes (~15s). Each episode (A and B) has regular oscillations corresponding to tremor before the end of freezing. The freezing episode showed in B also has some initial oscillatory movements that did not qualify as tremor according to the pre-established criteria. Also in B, the end of the freezing episode is followed by gait festination. The traces show raw accelerometry data. C and D, rate meters for the whole duration of the FOG episodes corresponding to A and B, respectively. The peaks correspond to the tremor periods towards the end of the freezing episode when walk restarts. Rate meters used the data produced after detection of full phase oscillations above the threshold. The graphs were constructed with a bin width of 500ms, and smoothed using a Gaussian filter. E, distribution of FOG tremor frequencies. The graph shows the frequencies (Hz) found across 20 recordings of tremor, each in a separate FOG episode. The average rate in the recorded tremors was 7.07Hz (±1.47 STD).
... Variability in the tremor associated with FOG in the MPTP-treated monkey. The oscillatory movements recorded with an accelerometer placed on the back of the leg during 3 episodes of tremor associated with FOG as examples of variability are shown in the three traces. The raster on top of each trace shows the detection of full phase oscillations above the threshold. The frequency and duration of the tremor were calculated on the constructed raster. Tremor frequencies in these episodes were: 6.0, 7.8, and 6.6Hz from top to bottom traces, respectively. Tremor durations in these episodes are shown next to each raster.
Contributors:Antonella Lombardi Costa, Cláubia Pereira, Walter Ambrosini, Francesco D’Auria
Relative power evolution during a period of oscillation of 1.06s.
Contributors:S.A. Galindo-Torres, A. Scheuermann, L. Li, D.M. Pedroso, D.J. Williams
Simulation setup. The no-flow boundary cells of the LBM domain were tagged as solid. The bounce back rule of Eq. (16) were applied to them but with cohesion (Eq. (17)) turned off to mitigate the effects of the boundaries. The water table oscillated with time during the simulation due to the varying pressure head imposed at the bottom boundary.
... Water saturation Sr as a function of dimensionless time (time multiplied by the factor g/H) from simulations with different frequencies (as indicated by the number of fluctuation cycles within the period of the plots).
... Amplitude of the Fourier components |F(ω)| for each frequency for both original (triangles) and filtered signals (squares).
... Amplitude A(ω) of the harmonic component of Sr(t) as a function of ω with a power law fitting (exponent equal to −0.36). Inset: phase angle ϕ representing the lag of the Sr oscillation behind the imposed head fluctuation.
... Imposed hydraulic head fluctuations (h0−h(t))/h0 versus the saturation variations (Sr) for two different frequencies and 4 cycles of equilibrium simulations. The arrows shows the time sequence taking for the equilibrium cycles, first imbibition and then drainage, which also holds for the dynamic simulations.
Contributors:M.M. Dernison, J.M.A.M. Kusters, P.H.J. Peters, W.P.M. van Meerwijk, D.L. Ypey, C.C.A.M. Gielen, E.J.J. van Zoelen, A.P.R. Theuvenet
Changes in intracellular Ca2+/Sr2+ levels of quiescent monolayer NRK cells, upon stimulation with PGF2α inside the ring, at different positions outside, inside and under the ring. Cells inside the ring were stimulation with PGF2α (n=7 experiments). Distribution of the interval time between calcium oscillations is depicted at right panels. (A) Response of 62 cells measured 5mm outside the ring at position A (see inset) after addition inside the ring of 0.8μM PGF2α. (B) Response of 47 cells measured under the ring (position B in inset) in the presence of PGF2α inside the ring. (C) Response of 67 cells measured at the inner edge of the ring (position C in inset) in the presence of PGF2α inside the ring. Individual traces are shown in gray, the mean response of the imaged cells is represented by the black line. Cells outside the ring were perfused with 5mM Sr2+, while inside the ring 1mM Ca2+ was present (see Materials and methods). The lower gray bar indicates presence of PGF2α inside the ring. Inset shows different positions of the imaged regions in relation to the ring location on the monolayer. Time (5min) is indicated by the lower black bar.
... The inset shows a schematic two-dimensional network of 400×400 cells. At the centre of the network cells, within a ring with an inner radius of 1mm (equivalent to 100 cells) have an IP3 concentration randomly distributed within a range between 5 and 15μM. (A) IP3 gradient due to diffusion and degradation of IP3 as described by Eq. (A.4) in Appendix A. For cells under the ring (0.5mm thick) the IP3 concentration decreases according to Eq. (A.4) from 10μM (inner side of the ring) to 0.1μM (outside the ring). Cells outside the ring have an IP3-concentration of 0.1μM and do not exhibit spontaneous calcium oscillations. (B) Membrane potential gradient in the network with (dashed line) and without (solid line) taking into account IP3 diffusion. The solid line shows the average membrane potential if there would be no diffusion of IP3 from the inside of the ring to cells under the ring. The dashed line shows the average membrane potential as a result of simulations for an IP3 distribution in the network according to Eq. (A.4) in Appendix A.
Contributors:L.D. Yépez, J.L. Carrillo, F. Donado, J.M. Sausedo-Solorio, P. Miranda-Romagnoli
The behavior of the average length of the clusters as a function of the frequency of one the fields, keeping the other frequency fixed (see text for more details).
... The average cluster length as a function of the frequency of the vertical rotating field.
... The mean cluster speed as a function of the frequency of the vertical rotating field.
... Plot of the dependence of the average chain-length as a function of the frequency of the rotating field. The effect of the viscous force limiting the cluster size is clearly seen even though the frequency is small. Obviously, the interval of frequency at which the average cluster length drops depends on the liquid viscosity.
... Average Mason number as a function of the frequency. Inset: Average Mason number as a function of the phase shift.
Contributors:Sandra J. Niederschuh, Hartmut Witte, Manuela Schmidt
Forelimb (“limb”) stride frequency and whisking (“whisk”) frequency. Range, mean value±SD, and speed dependence (F-value and coefficient of determination r2 in %).
... Variation of frequency across the speed range on the continuous (con) and discontinuous (dis) substrate in the presence (vib pres) or absence (carp abs) of carpal vibrissae. (A) Forelimb stride frequency. (B) Whisking frequency.
... Temporal and spatial parameters of forelimb kinematics compared across all subsets of data: all vibrissae present (vp), carpal vibrissae absent (ca), mystacial vibrissae absent (ma), and all vibrissae absent (va). Box plots illustrate the median (bold line), the range between the upper and the lower quartile (box) and maximum and minimum values. (A) Forelimb stride frequency. (B) Forepaw placement relative to the anterior margin of the orbita (=X0). (C) Limb angle at touchdown.
Contributors:William F. Louisos, Darren L. Hitt, Christopher M. Danforth
The temporal evolution of mass flow rate in the thermosyphon at a Rayleigh number of Ra∼7.08×105. The flow undergoes four initial, transient flow reversals before settling into a state of growing, CCW oscillations which then experience flow reversals after the oscillations have grown sufficiently large.
... The temporal evolution of mass flow rate in the thermosyphon at a Rayleigh number of Ra∼2.83×107. The flow undergoes several initial, transient flow reversals before establishing a steady, high-Rayleigh, aperiodic, stable convective flow regime with small amplitude, high frequencyoscillations centered on a mass flow RMS of approximately 0.525kg/s.
... The temporal evolution of mass flow rate in the thermosyphon at a Rayleigh number of Ra∼2.83×104. This flow regime is characterized as damped, stable convection as the oscillations remain CW and decay to the RMS, steady-state value.
... The dominant oscillatory frequency of the mass flow pulsation magnitude in the thermosyphon. Dominant frequencies are determined by the peak in the power spectra from the Fourier transform of the temporal mass flow evolution. It is found that the dominant frequency scales as Ra0.48.
... The temporal evolution of mass flow rate in the thermosyphon at a Rayleigh number of Ra∼1.42×106. The flow undergoes initial, transient flow reversals during start-up, oscillations then begin small and grow large until a flow reversal occurs and a fully chaotic flow regime is established.