led light stimulation (470  (Thorlabs)

Journal: Nature

Article Title: Dendritic, delayed, stochastic CaMKII activation in behavioural time scale plasticity

doi: 10.1038/s41586-024-08021-8

Figure Lengend Snippet: a , Experimental set up (top), BTSP protocol (middle) and raw traces of EPSPs (ten recordings and average) before and after BTSP protocol (bottom). b , Fluorescence lifetime images (top) and time course (bottom) of a dendritic segment expressing 2dV-Camui during BTSP induction (shaded region, BTSP protocol). c , Representative traces (top) and all dendrites (bottom) for which fluorescence lifetime changes of 2dV-Camui were measured after BTSP (left) and no stimulation (no stim, right). Shaded region, BTSP protocol. d , Frequency of DDSC onsets after BTSP and cBTSP induction compared with uncaging only (uncage), current injection only (depol) and no stimulation controls. e , Cumulative frequency of d . Two-sided Kolmogorov–Smirnov test with Bonferroni’s correction, P = 0.00032 for BTSP versus uncage; P = 0.0000072 for BTSP versus depol; P = 0.00000026 for BTSP versus no stim; and P = 0.12 for BTSP versus cBTSP. f , g , Averaged time course ( f ) and area under the curve (30–40 s after BTSP) ( g ) of lifetime change of CaMKII activity under all five conditions. Data are presented as the mean ± s.e.m. in f and median and second quartile along with individual values in g . Two-way ANOVA ( F 4,100 = 5.434, P = 0.0005) with Dunnett’s multiple comparison test (all P values are shown in the source data file). NS, not significant. h , Fluorescence lifetime images of 2dV lifetime changes before (−2 s) and after (+50 s) BTSP protocol at secondary dendrites or soma. i , Representative 2dV-Camui traces of dendrite and of soma or primary dendrites from eight different cells. If the soma was not in the same z plane as the stimulated dendrite, the primary dendrite was used. j , Pie chart showing percentage of recordings that showed an increase in CaMKII activity either in both stimulated dendrite and soma (or primary dendrite) or specifically in the dendrites. k , Amplitude of DDSC in BTSP-induced dendrites and in soma or primary dendrites. The data are presented as the mean and individual values. Two-tailed paired t -test ( t 19 = 4.924, P = 0.000094). Plus symbol in images depicts the BTSP-stimulated spine. Number of dendrites are mentioned in the figure panels in parentheses wherever appropriate. * P < 0.05, *** P < 0.001. Scale bars, 2.5 μm ( b ) or 5 μm ( h ).

Article Snippet: LED light stimulation (470 nm, M470L5, Thorlabs) was used to activate paAIP2.

Techniques: Fluorescence, Expressing, Injection, Activity Assay, Comparison, Two Tailed Test

Journal: Nature

Article Title: Dendritic, delayed, stochastic CaMKII activation in behavioural time scale plasticity

doi: 10.1038/s41586-024-08021-8

Figure Lengend Snippet: a , Left: Representative dendritic 2dV-Camui recordings in basal dendrites before induction of BTSP. Right: All 2dV-Camui recordings (filtered) before BTSP induction. b , Same as (a) , but performed after BTSP induction (shaded region, BTSP protocol). c , Same as (a) , but performed in corresponding proximal apical dendrites after BTSP induction (shaded region, BTSP protocol). d , Frequency of DDSC onsets after BTSP induction in apical (orange) and basal dendrites (red) compared with no stimulation (black) controls. e , Cumulative frequency of (d) . Two-sided Kolmogorov-Smirnov test with Boenhoffer’s correction, basal vs apical p = 1.0; basal vs no stim p = 0.018. The number of dendrites is mentioned in the figure panels. *p < 0.05.

Article Snippet: LED light stimulation (470 nm, M470L5, Thorlabs) was used to activate paAIP2.

Techniques:

Journal: Nature

Article Title: Dendritic, delayed, stochastic CaMKII activation in behavioural time scale plasticity

doi: 10.1038/s41586-024-08021-8

Figure Lengend Snippet: a , Left, schematic of the photoactivatable CaMKII inhibitor paAIP2, which inhibits CaMKII after blue light (BL, 470 nm) exposure and becomes inactive within about 40 s when BL is stopped. Right, paAIP2-P2A-mEGFP-labelled CA1 neurons. Scale bar, 50 μm. b , Schematic of two separate CaMKII inhibition experiments shown in c , e and f . CaMKII was inhibited for 2 mins, 0 s (black) or 30 s (blue) after the BTSP protocol (orange). c , Representative EPSP traces of a stimulated spine (average of ten recordings) before and after BTSP induction in paAIP2-labelled neurons for which no BL stimulation was given (orange), or BL was given with 0 s delay (black) or 30 s delay (blue). d – g , Normalized time course ( d – f ) and summary of magnitude of EPSP potentiation (25–30 min) ( g ) in stimulated and adjacent spines for no BL ( n = 17; d ), BL with 0 s delay ( n = 10; e ), BL with 15 s delay ( n = 10) or BL with 30 s delay ( n = 11; f ). Two way ANOVA ( F 7,68 = 4.94, P = 0.0001) with Tukey’s multiple comparison test (all P values are shown in the source data file). h , Top, BTSP protocol in voltage clamp (BTSPvc), for which 5 uncaging pulses (1 Hz) were paired with depolarization to 0 mV for 300 ms with 750 ms delay in the presence of TTX. Middle, protocol to artificially induce delayed CaMKII activity by applying additional long (20 s) depolarization 10–12 s after BTSPvc (BTSPvc+depol). Bottom, 5 uncaging pulses were paired with long depolarization 10–12 s after uncaging (uncage+depol). i , Representative EPSC traces of stimulated spine (average of 10 recordings) before and 20 min after BTSPvc, BTSPvc+depol and uncage+depol. j , k , Normalized EPSC amplitude time course ( j ) and summary of magnitude of potentiation ( k ) of stimulated and adjacent spines during BTSPvc ( n = 10), BTSPvc+depol (stim, n = 19; adj, n = 11) and uncage+depol (stim, n = 14; adj, n = 13). Two-way ANOVA ( F 5,53 = 4.9, P = 0.0009) with Tukey’s multiple comparison test (all P values are shown in the source data file). The data in d – g , j and k are presented as the mean ± s.e.m. Number of cells are mentioned in the panels in parentheses wherever appropriate. Arrows depict BTSP, BTSPvc or uncaging induction. * P < 0.05, *** P < 0.001.

Article Snippet: LED light stimulation (470 nm, M470L5, Thorlabs) was used to activate paAIP2.

Techniques: Inhibition, Comparison, Activity Assay