Journal: The Journal of Physiology

Article Title: Glial EAAT2 regulation of extracellular nTS glutamate critically controls neuronal activity and cardiorespiratory reflexes

doi: 10.1113/JP274620

Figure Lengend Snippet: A, representative example showing general iGluR blockade with Kyn (40 mm, bars, n = 9) in the nTS increased arterial pressure (AP) and splanchnic sympathetic nerve activity (SSNA) with variable changes in phrenic (Phr) nerve activity. Subsequent responses to DHK (post Kyn, bars, 2 mm) were diminished compared to those in the absence of blockade (see Fig. 3). B–G, Kyn attenuated responses to DHK (normalized to control DHK responses, 100%, dashed line): mean arterial pressure (MAP, B), heart rate (HR, C), splanchnic sympathetic nerve activity (SSNA, D), and phrenic (Phr) frequency (Freq, E), amplitude (Amp, F) and minute activity (Min PhrNA, G). The AMPA receptor antagonist NBQX (2 mm, n = 13) similarly inhibited DHK responses. In contrast, the NMDA receptor antagonist AP5 (10 mm, n = 16) augmented the effects of DHK on MAP, SSNA, Phr Freq and also Min PhrNA (P = 0.07). Paired t test, relative to respective control DHK responses: * P < 0.05, ** P < 0.01, *** P < 0.001. One‐way ANOVA, relative to the effects of Kyn or NBQX: †† P < 0.01, ††† P < 0.001.

Article Snippet: Reagents DHK, NBQX (AMPAR blocker) and AP5 (NMDAR blocker) were purchased from Tocris Bioscience (R&D Systems, Inc., Minneapolis, MN, USA).

Techniques: Activity Assay, Control

Journal: The Journal of Physiology

Article Title: Glial EAAT2 regulation of extracellular nTS glutamate critically controls neuronal activity and cardiorespiratory reflexes

doi: 10.1113/JP274620

Figure Lengend Snippet: A–D, representative examples of DHK‐induced currents in the absence and presence of glutamate receptor block. A, prior application of Kyn blocks DHK‐currents. B–D, following an initial DHK‐induced inward current, a second application of DHK was made in the presence of NBQX and AP5 (n = 6; B), NBQX (n = 8; C), or AP5 (n = 8; D). E, magnitude of I hold changes by DHK in the presence of iGluR antagonists, relative to the first DHK‐induced current (defined as 100%). Note the attenuation of DHK‐currents in the presence of NBQX and NBQX + AP5. One‐way RM ANOVA, relative to respective control DHK responses: * P < 0.05, ** P < 0.01.

Article Snippet: Reagents DHK, NBQX (AMPAR blocker) and AP5 (NMDAR blocker) were purchased from Tocris Bioscience (R&D Systems, Inc., Minneapolis, MN, USA).

Techniques: Blocking Assay, Control

Journal: bioRxiv

Article Title: Graded heterogeneity of metabotropic signaling underlies a continuum of cell-intrinsic temporal responses

doi: 10.1101/2020.12.27.424473

Figure Lengend Snippet: a. Schematics of cerebellar circuit b. UMAP embedding showing normalized expression of gene involved in mGluR1 signaling cascade. c. Same as in D for AMPA and NMDA receptors. d. Same as in D for genes involved in mGluR2 signaling cascade. e. Example spiking responses in different UBCs to a burst of MF input (20 stimuli at 100Hz). f. Instantaneous firing rate for the same cells.

Article Snippet: Sequential drug wash-ins were performed using a computer-controlled solenoid manifold system (ValveLink 8.2, Automate Scientific, Inc., Berkeley, CA) with a flow rate of 1 to 2mL/min, where indicated, with mGluR1 antagonist (100μM LY357385, Tocris Bio-Techne, Minneapolis, MN), AMPA receptor blocker (5μM NBQX), NMDA receptor blocker (2 μM R-CPP), mGluR2/3 antagonist (1μM LY341495, Tocris Bio-Techne, Minneapolis, MN) and DGK inhibitor II (100 μM R59949, MilliporeSigma, St. Louis, MO)

Techniques: Expressing

Journal: bioRxiv

Article Title: Graded heterogeneity of metabotropic signaling underlies a continuum of cell-intrinsic temporal responses

doi: 10.1101/2020.12.27.424473

Figure Lengend Snippet: a. Examples of instantaneous firing rate from on-cell recordings before (black) and after (gray) the application of AMPA/NMDA receptor antagonist (gray bar) in a fast (left) and a slow (right) UBC. Each trace is an average of 8 trials. b. Summary of evoked spiking (20×100 Hz) and the effect of AMPA/NMDA receptor antagonist (normalized to baseline, mean±sem, n=5). c. Examples of instantaneous firing rate from on-cell recordings before (black) and after (red) the application of an mGluR1 antagonist (red bar) in a fast (left) and a slow (right) UBC. Each trace is an average of 8 trials. d. Summary of evoked spiking (20×100 Hz) and the effect of an mGluR1 antagonist (normalized to baseline, mean±sem, n=5). e. Example of instantaneous firing rate before and after 20 minutes of DGK inhibitor II wash-in. f. Summary of half-decay time of instantaneous firing rate response with DGK inhibitor II (normalized to baseline, mean±sem, n=6) g. Summary of peak amplitude of instantaneous firing rate response with DGK inhibitor II (normalized to baseline, mean±sem, n=6)

Article Snippet: Sequential drug wash-ins were performed using a computer-controlled solenoid manifold system (ValveLink 8.2, Automate Scientific, Inc., Berkeley, CA) with a flow rate of 1 to 2mL/min, where indicated, with mGluR1 antagonist (100μM LY357385, Tocris Bio-Techne, Minneapolis, MN), AMPA receptor blocker (5μM NBQX), NMDA receptor blocker (2 μM R-CPP), mGluR2/3 antagonist (1μM LY341495, Tocris Bio-Techne, Minneapolis, MN) and DGK inhibitor II (100 μM R59949, MilliporeSigma, St. Louis, MO)

Techniques:

Journal: bioRxiv

Article Title: Graded heterogeneity of metabotropic signaling underlies a continuum of cell-intrinsic temporal responses

doi: 10.1101/2020.12.27.424473

Figure Lengend Snippet: a. Sample cell-attached recording (top) instantaneous firing rate (middle) and synaptic current measured with whole-cell voltage clamp (bottom) in a cell with fast response. b. Same as in A but for a cell with intermediate speed response c. Same as in A but for a cell with clear biphasic synaptic current (bottom) d. Same as in A but for a cell with slow biphasic response e. Same as in A but for a cell with only a pause in firing f. Half-decay times of firing rates vs. half-decay times of currents and linear fit on a log-log plot (R adj 2 = 0.91, slope = 0.88, intercept = -0.22, n = 17) g. Peak firing rate vs. peak current amplitude and linear fit on a log-log plot (R adj 2 = 0.69, slope = 0.71, intercept = -0.65, n = 17) h. Pause duration vs. amplitude of the current at stimulation offset, and linear fit with log 10 response variable (black, R adj 2 = 0.44, slope = 0.01, intercept = -0.42, n = 20) i. Heatmap of peak normalized mGluR1-mediated current (n=10) j. Heatmap of peak normalized mGluR2/3-mediated current (n=7) k. Average synaptic response before (black), after (red) mGluR1 antagonist, and after AMPA/NMDA receptor antagonist wash-ins (grey). Each trace is an average of 8 trials. l. Average synaptic response before (top, black) and after (top, blue) mGluR2/3 antagonist washin, and their difference (bottom, black). Each trace is an average of 8 trials.

Article Snippet: Sequential drug wash-ins were performed using a computer-controlled solenoid manifold system (ValveLink 8.2, Automate Scientific, Inc., Berkeley, CA) with a flow rate of 1 to 2mL/min, where indicated, with mGluR1 antagonist (100μM LY357385, Tocris Bio-Techne, Minneapolis, MN), AMPA receptor blocker (5μM NBQX), NMDA receptor blocker (2 μM R-CPP), mGluR2/3 antagonist (1μM LY341495, Tocris Bio-Techne, Minneapolis, MN) and DGK inhibitor II (100 μM R59949, MilliporeSigma, St. Louis, MO)

Techniques: