Journal: iScience

Article Title: The endoplasmic reticulum plays a key role in α-cell intracellular Ca 2+ dynamics and glucose-regulated glucagon secretion in mouse islets

doi: 10.1016/j.isci.2024.109665

Figure Lengend Snippet: L-type Ca v -channels carry the bulk of transmembrane Ca 2+ and are required for α-cell electrical activity, but not glucagon secretion (A) Depolarisation-triggered Ca 2+ current recorded in an α-cell, in response to sequential addition of tetrodotoxin (TTX; Na v blocker), isradipine (israd; Red; L-type Ca v channel blocker), ω-agatoxin IVA (ATX; blue; P/Q-type Ca v channel blocker), and ω-conotoxin GIVA (CTX; Green; N-type Ca v channel blocker) as indicated. The shaded areas with respective colors mark the proportion of Ca 2+ current blocked by the specific blockers. (B) Relative contribution of Ca v channel channels to transmembrane Ca 2+ current, as measured in (A). L-type Ca v channel, L; P/Q-type Ca v channel, P/Q; N-type Ca v channel, N. (C) Membrane potential recording of an α-cell in the presence of 1 mM glucose. Application of 2 μM isradipine and 10 μM BayK8644 are marked by horizontal bars over the trace. Representative responses under control (❶), 2 μM isardipine (❷) and 10 μM BayK8644 (❸) are displayed below on expanded timescale (bottom). (D) Glucagon secretion from batch-incubated islets in response to 1 and 6 mM glucose in the absence (black bars, n = 11) or presence of 2 μM ( n = 4) or 10 μM ( n = 11) isradipine (red bars) alone or in combination with 200 nM ω-agatoxin IVA (purple bars; n = 7). Data are normalized to secretion at 1 mM glucose in the control and presented as mean values ±SEM. ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗∗ p < 0.0001 for indicated comparisons.

Article Snippet: BayK8644 , Tocris , Cat# 1544.

Techniques: Activity Assay, Membrane, Control, Incubation

Journal: iScience

Article Title: The endoplasmic reticulum plays a key role in α-cell intracellular Ca 2+ dynamics and glucose-regulated glucagon secretion in mouse islets

doi: 10.1016/j.isci.2024.109665

Figure Lengend Snippet:

Article Snippet: BayK8644 , Tocris , Cat# 1544.

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Plasmid Preparation, Software

Journal: bioRxiv

Article Title: Calcium dynamics tune developmental tempo to generate evolutionarily divergent axon tract lengths

doi: 10.1101/2024.12.28.630576

Figure Lengend Snippet: a. Axon tract of human (day 70) brain organoid slices in micropatterned devices expressing GCaMP7f and treated with DMSO (control), nimodipine (L-Type VGCC antagonist) or BayK8644 (L-Type VGCC agonist). Arrowheads mark axons showing calcium activity in a five minute timelapse, indicating nearly complete perturbation of calcium dynamics upon L-Type VGCC blockage. Zooms show representative timelapse of individual axons, with ΔF/F0 fluorescent GCaMP7f intensity plots of axons in zooms, showing the increase in calcium dynamics upon L-Type VGCC activation. b. Quantifications of number of axons firing in an ROI during a five minutes timelapse in axon tracts of human brain organoid slices (day 70) in micropatterned devices expressing GCaMP7f and treated with DMSO (control), nimodipine (L-Type VGCC antagonist) or BayK8644 (L-Type VGCC agonist). Control: n=13 regions of interest, L-Type VGCC antagonist: n=11, L-Type VGCC agonist: n=10. c. Quantifications of average mean amplitudes of spontaneous calcium transients in human brain organoid slice cultures (day 70) in micropatterned devices expressing GCaMP7f and treated with DMSO (control) or BayK8644 (L-Type VGCC agonist). Control: n=137 axons, L-Type VGCC agonist: n=100. d. Quantifications of average frequencies of spontaneous calcium transients in human brain organoid slice cultures (day 70) in micropatterned devices expressing GCaMP7f and treated with DMSO (control) or BayK8644 (L-Type VGCC agonist). Control: n=137 axons, L-Type VGCC agonist: n=100. e. Quantifications of average event durations of spontaneous calcium transients in human brain organoid slice cultures (day 70) in micropatterned devices expressing GCaMP7f and treated with DMSO (control) or BayK8644 (L-Type VGCC agonist). Control: n=137 axons, L-Type VGCC agonist: n=100. f. Representative images of axon tracts of human (day 90) brain organoid slices in micropatterned devices immunostained for pan-axonal neurofilaments and treated with DMSO (control), nimodipine (L-Type VGCC antagonist), BayK8644 (L-Type VGCC agonist), or cAMP. g. Quantifications of axon tract length of human brain organoid slice cultures in micropatterned devices, treated with DMSO (control), nimodipine (L-Type VGCC antagonist) or BayK8644 (L-Type VGCC agonist), at different timepoints. n=8-32 organoids. h. Quantifications of axon tract length of human brain organoid slice cultures in micropatterned devices, treated with DMSO (control) or BayK8644 (L-Type VGCC agonist), at day 55+7 and 55+28. Control: n=15 organoids (55+7) and n=14 (55+28), L-Type VGCC agonist: n=11 (55+7) and n=8 (55+28). i. Quantifications of axon tract length of human brain organoid slice cultures in micropatterned devices, treated with DMSO (control) or cAMP, at different timepoints. n=4-5 organoids. j. Quantifications of axon tract length of human brain organoid slice cultures in micropatterned devices, treated with DMSO (control) or cAMP, at day 55+7 and 55+28. Control: n=10 organoids (55+7) and n=10 (55+28), cAMP: n=9 (55+7) and n=9 (55+28). k. Schematic of model. Increase of L-type VGCC-mediated calcium, and downstream cAMP, levels accelerate developmental tempo during axonogenesis, resulting in shorter axon tract lengths. treatm., treatment. antag., antagonist. ag., agonist. All data are shown as mean ± SEM; ** P < 0.1, **** P < 0.0001 as determined by unpaired t tests.

Article Snippet: For drug treatments, one day after brain organoid slices were placed in the devices, and during subsequent medium changes, medium was replaced with SFSCM or BrainPhys (at similar timepoints as with ALI-CO cultures described above) supplemented with indicated final concentrations of either 1 μM or 10 μM BayK8644 (Stratech Scientific, S7924-SEL), 1 μM or 10 μM Nimodipine (Selleckchem, S1747) or 100 μM 8-Br-cAMP (BioLog, B007).

Techniques: Expressing, Control, Activity Assay, Activation Assay

Journal: Frontiers in Cardiovascular Medicine

Article Title: Unveiling the Role of DNA Methylation in Vascular CACNA1C Tissue–Specific Expression

doi: 10.3389/fcvm.2022.872977

Figure Lengend Snippet: BayK8644-induced vasoconstrictions and [Ca 2+ ]i signals. (A–C) BayK8644-induced Ca 2+ increases in single isolated vascular SMC from TA, MA, MCA, or PA. Representative traces (A) and statistics (B) of Ca 2+ transients elicited by BayK8644 (10 −5 mol/L) measured with the fluorescence Ca 2+ indicator Fluo-3 AM. Maximum of Ca 2+ responses (C) were calculated. The fractional fluorescence intensity was calculated as F/F0, where F is the fluorescence intensity for the region of interest, and F0 is the fluorescence intensity during a period from the beginning of the recording when there was no Ca 2+ activity (n =30 cells from ten rat per group), (D) Representative tracings of BayK8644-induced vasoconstrictions in TA, MA, MCA, or PA rings, (E) Statistics of BayK8644-induced vasoconstrictions and pD2 in vessel rings ( N = 4, n = 8). N, number of adult male offspring; n, number of vessel rings. pD2, –log[50% effective concentration]. Data were presented as means ± SEM.

Article Snippet: The vessel rings were contracted with cumulatively increasing concentrations of BayK8644 (an agonist for voltage-dependent Ca 2+ channels; 10 −10 -10 −5 mol/L; Sigma-aldrich, #B112).

Techniques: Isolation, Fluorescence, Activity Assay, Concentration Assay