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<t>NGF</t> attenuates <t>CAPS</t> tachyphylaxis. Response ratios (peak value obtained for second CAPS stimulation after [A]/peak value for first stimulation before [B] exposure to 100 ng/mL NGF) for the ( A ) number of responders or ( B – D ) Max. signal intensity evoked during the second exposure to 0.3 (blue) or 1 μM CAPS (red) divided by the values elicited the first time when the DRG were stimulated with the same concentration of agonist. In ( B – D ), the calculations were restricted to cells that conformed to the following criteria: ( B ) neurons that produced above threshold signals to 0.3 or 1 μM CAPS the first time each concentration was applied; note that both above and below threshold values elicited by the second stimulation with either agonist concentration were included in these analyses, ( C ) neurons excited above threshold both times the DRG were exposed to 0.3 or 1 μM CAPS, ( D ) all DRGNs excited at least one time by either the first or second stimulation with 0.3 or 1 μM CAPS; although the threshold criterion was used to identify the neurons to analyse, comparisons before and after NGF included values both above and below threshold. Paired two-tailed Student’s t-test were performed to compare the mean values of [A] and [B]; * p

Ngf, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "Ca2+ Signalling Induced by NGF Identifies a Subset of Capsaicin-Excitable Neurons Displaying Enhanced Chemo-Nociception in Dorsal Root Ganglion Explants from Adult pirt-GCaMP3 Mouse"

Article Title: Ca2+ Signalling Induced by NGF Identifies a Subset of Capsaicin-Excitable Neurons Displaying Enhanced Chemo-Nociception in Dorsal Root Ganglion Explants from Adult pirt-GCaMP3 Mouse

Journal: International Journal of Molecular Sciences

doi: 10.3390/ijms22052589

NGF attenuates CAPS tachyphylaxis. Response ratios (peak value obtained for second CAPS stimulation after [A]/peak value for first stimulation before [B] exposure to 100 ng/mL NGF) for the ( A ) number of responders or ( B – D ) Max. signal intensity evoked during the second exposure to 0.3 (blue) or 1 μM CAPS (red) divided by the values elicited the first time when the DRG were stimulated with the same concentration of agonist. In ( B – D ), the calculations were restricted to cells that conformed to the following criteria: ( B ) neurons that produced above threshold signals to 0.3 or 1 μM CAPS the first time each concentration was applied; note that both above and below threshold values elicited by the second stimulation with either agonist concentration were included in these analyses, ( C ) neurons excited above threshold both times the DRG were exposed to 0.3 or 1 μM CAPS, ( D ) all DRGNs excited at least one time by either the first or second stimulation with 0.3 or 1 μM CAPS; although the threshold criterion was used to identify the neurons to analyse, comparisons before and after NGF included values both above and below threshold. Paired two-tailed Student’s t-test were performed to compare the mean values of [A] and [B]; * p

Figure Legend Snippet: NGF attenuates CAPS tachyphylaxis. Response ratios (peak value obtained for second CAPS stimulation after [A]/peak value for first stimulation before [B] exposure to 100 ng/mL NGF) for the ( A ) number of responders or ( B – D ) Max. signal intensity evoked during the second exposure to 0.3 (blue) or 1 μM CAPS (red) divided by the values elicited the first time when the DRG were stimulated with the same concentration of agonist. In ( B – D ), the calculations were restricted to cells that conformed to the following criteria: ( B ) neurons that produced above threshold signals to 0.3 or 1 μM CAPS the first time each concentration was applied; note that both above and below threshold values elicited by the second stimulation with either agonist concentration were included in these analyses, ( C ) neurons excited above threshold both times the DRG were exposed to 0.3 or 1 μM CAPS, ( D ) all DRGNs excited at least one time by either the first or second stimulation with 0.3 or 1 μM CAPS; although the threshold criterion was used to identify the neurons to analyse, comparisons before and after NGF included values both above and below threshold. Paired two-tailed Student’s t-test were performed to compare the mean values of [A] and [B]; * p

Techniques Used: Concentration Assay, Produced, Two Tailed Test

Brief exposure to NGF results in sensitisation to CAPS. Analysis tools were applied to signals evoked by 0.3 (blue dots) and 1 μM CAPS (red dots) before [B] and after [A] 5 min exposure to 100 ng/mL NGF (blue trace in Figure 1 A) to measure ( A ) Max. intensity, ( B ) signal duration and ( C ) lag time. Asterisks represent significance between the measurements before and after NGF treatment, determined by Student’s t-test for unpaired samples, unequal variance; * p

Figure Legend Snippet: Brief exposure to NGF results in sensitisation to CAPS. Analysis tools were applied to signals evoked by 0.3 (blue dots) and 1 μM CAPS (red dots) before [B] and after [A] 5 min exposure to 100 ng/mL NGF (blue trace in Figure 1 A) to measure ( A ) Max. intensity, ( B ) signal duration and ( C ) lag time. Asterisks represent significance between the measurements before and after NGF treatment, determined by Student’s t-test for unpaired samples, unequal variance; * p

Techniques Used:

$NGF evokes signals if applied before CAPS and excites a new cohort when re-applied afterwards; CAPS-induced signal strength correlates with responsiveness to NGF. ( A ) Summed increase in fluorescence from DRG treated, in the following order, with 100 ng/mL NGF (20 min followed by 40 min washout, [N1]); 1 μM CAPS (5 min, 15 min washout, [C1]); NGF for a second time (20 min, 20 min washout, [N2]); 1 μM CAPS again (5 min, 15 min washout, [C2]) and, finally, 10 μM CAPS (5 min, 35 min washout, [C3]). ( B ) Shows the relative increase in the number of active cells (number activated by 10 μM CAPS [C3]/number excited by 1 μM [C1]) calculated for all CAPS-excitable cells (Total) or only the subsets that were NGF-excitable and -refractory cells. ( C – E ) Neurons that responded during the first ([C1]; blue dots) or second exposure ([C2]; green dots) to 1 μM CAPS and those excited by 10 μM CAPS ([C3]; red dots) were sub-categorised according to whether they were excited both times (2, abscissa) on exposure to NGF (i.e., during [N1] and [N2]), only once (1, abscissa) by either application (i.e., [N1] or [N2]) or remained inactive both times (0, abscissa). Each sub-category was analysed for ( C ) lag time, ( D ) signal duration and ( E ) Max. intensity. Plotted data represents mean ± s.e.m. Asterisks indicate p values for Student’s t -tests compared between group 1 and the requisite data set in group 0, or between groups 1 and 2. ** p$
Figure Legend Snippet: NGF evokes signals if applied before CAPS and excites a new cohort when re-applied afterwards; CAPS-induced signal strength correlates with responsiveness to NGF. ( A ) Summed increase in fluorescence from DRG treated, in the following order, with 100 ng/mL NGF (20 min followed by 40 min washout, [N1]); 1 μM CAPS (5 min, 15 min washout, [C1]); NGF for a second time (20 min, 20 min washout, [N2]); 1 μM CAPS again (5 min, 15 min washout, [C2]) and, finally, 10 μM CAPS (5 min, 35 min washout, [C3]). ( B ) Shows the relative increase in the number of active cells (number activated by 10 μM CAPS [C3]/number excited by 1 μM [C1]) calculated for all CAPS-excitable cells (Total) or only the subsets that were NGF-excitable and -refractory cells. ( C – E ) Neurons that responded during the first ([C1]; blue dots) or second exposure ([C2]; green dots) to 1 μM CAPS and those excited by 10 μM CAPS ([C3]; red dots) were sub-categorised according to whether they were excited both times (2, abscissa) on exposure to NGF (i.e., during [N1] and [N2]), only once (1, abscissa) by either application (i.e., [N1] or [N2]) or remained inactive both times (0, abscissa). Each sub-category was analysed for ( C ) lag time, ( D ) signal duration and ( E ) Max. intensity. Plotted data represents mean ± s.e.m. Asterisks indicate p values for Student’s t -tests compared between group 1 and the requisite data set in group 0, or between groups 1 and 2. ** p

Techniques Used: Fluorescence

NGF evokes Ca2+ signals in a subset of CAPS-excitable cells. ( A ) Traces show increases in fluorescence intensity (summed from 3 experiments) in DRGNs exposed sequentially to CAPS for 5 min. (black bars, at concentrations indicated) and 100 ng/mL NGF for 5 (blue) or 20 min. (red). In the latter case, a 5 min. washout after the 20 min. exposure to NGF has been omitted from the figure to keep the subsequent responses to CAPS in register with those in the blue trace. Note that F 0 was re-zeroed (see Section 4 ) before each addition of CAPS or NGF. Examples of fluorescence traces from individual DRGNs are presented in Supplementary Figure S1 . ( B ) Shows the mean ± s.e.m. Max. fluorescence intensity increases in neurons that responded above threshold upon exposure to each stimulus, as indicated on abscissa, in DRG exposed to NGF for 5 (blue bars) or 20 min. (red bars). Significant differences were observed for 0.3 μM CAPS before NGF ( p = 0.03), reflecting minor variation between experimental groups, and for 1 μM CAPS after NGF ( p = 0.02) but this latter difference might be due to the high background fluorescence after 20 min. with NGF, which may interfere with the quantification of subsequent responses to CAPS. ( C ) Number of active cells counted in 1 min. intervals after the addition of NGF, plotted against time; data summed from 3 independent recordings for each NGF treatment. ( D ) Mean number of cells activated (± s.e.m., N = 3) over 20 min. after the addition of CAPS (at concentrations indicated), or NGF, in DRG exposed to the latter for 5 (blue) or 20 min. (red). Results obtained with either CAPS concentration are shown for a two-tailed Student’s t -test between groups exposed to NGF for 5 min. and those treated for 20 min; * p

Figure Legend Snippet: NGF evokes Ca2+ signals in a subset of CAPS-excitable cells. ( A ) Traces show increases in fluorescence intensity (summed from 3 experiments) in DRGNs exposed sequentially to CAPS for 5 min. (black bars, at concentrations indicated) and 100 ng/mL NGF for 5 (blue) or 20 min. (red). In the latter case, a 5 min. washout after the 20 min. exposure to NGF has been omitted from the figure to keep the subsequent responses to CAPS in register with those in the blue trace. Note that F 0 was re-zeroed (see Section 4 ) before each addition of CAPS or NGF. Examples of fluorescence traces from individual DRGNs are presented in Supplementary Figure S1 . ( B ) Shows the mean ± s.e.m. Max. fluorescence intensity increases in neurons that responded above threshold upon exposure to each stimulus, as indicated on abscissa, in DRG exposed to NGF for 5 (blue bars) or 20 min. (red bars). Significant differences were observed for 0.3 μM CAPS before NGF ( p = 0.03), reflecting minor variation between experimental groups, and for 1 μM CAPS after NGF ( p = 0.02) but this latter difference might be due to the high background fluorescence after 20 min. with NGF, which may interfere with the quantification of subsequent responses to CAPS. ( C ) Number of active cells counted in 1 min. intervals after the addition of NGF, plotted against time; data summed from 3 independent recordings for each NGF treatment. ( D ) Mean number of cells activated (± s.e.m., N = 3) over 20 min. after the addition of CAPS (at concentrations indicated), or NGF, in DRG exposed to the latter for 5 (blue) or 20 min. (red). Results obtained with either CAPS concentration are shown for a two-tailed Student’s t -test between groups exposed to NGF for 5 min. and those treated for 20 min; * p

Techniques Used: Fluorescence, Concentration Assay, Two Tailed Test

Acute sensitisation by NGF of responses to CAPS is most prevalent in cells that exhibit [Ca2+]i signals when the neurotrophin is applied. Neurons that responded to 1 μM CAPS both times when it was applied before and after 5 min exposure to NGF (blue trace in Figure 1 A) were split into two groups depending on whether they additionally exhibited Ca 2+ signals in response to NGF alone (red dots, n = 13 neurons from 3 DRG recordings) or did not (blue dots, n = 170). The analysis tools were applied to measure ( A ) Max. intensity, ( B ) signal duration and ( C ) lag time for the CAPS-evoked signals before [B] (on abscissa) and after [A] the treatment with NGF. Asterisks show significant differences within groups between responses before and after NGF treatment (Student’s t -test, paired samples; * p

Figure Legend Snippet: Acute sensitisation by NGF of responses to CAPS is most prevalent in cells that exhibit [Ca2+]i signals when the neurotrophin is applied. Neurons that responded to 1 μM CAPS both times when it was applied before and after 5 min exposure to NGF (blue trace in Figure 1 A) were split into two groups depending on whether they additionally exhibited Ca 2+ signals in response to NGF alone (red dots, n = 13 neurons from 3 DRG recordings) or did not (blue dots, n = 170). The analysis tools were applied to measure ( A ) Max. intensity, ( B ) signal duration and ( C ) lag time for the CAPS-evoked signals before [B] (on abscissa) and after [A] the treatment with NGF. Asterisks show significant differences within groups between responses before and after NGF treatment (Student’s t -test, paired samples; * p

Techniques Used:

$NGF-excitable cells exhibit more robust responses to CAPS. Further analysis was performed on DRG subjected to the protocol described for the red trace in Figure 1 A. Cells were categorised according to whether or not they responded to 20 min. with NGF. ( A ) The number of NGF-excitable (red bars) and -refractory (blue bars) cells that responded to each treatment with the indicated CAPS concentrations applied before [B] or after [A] NGF. Analysis tools were applied to each sub-category to measure ( B ) Lag, ( C ) Duration and ( D ) Max. intensity, in all cases plotted as means ± s.e.m., with blue and red dots representing, respectively, values from individual cells assigned to the NGF-refractory and -excitable categories. Black asterisks represent p values derived by unpaired Student’s t-test between the NGF-excitable and –refractory cells within each CAPS stimulation group. Coloured asterisks show results from paired Student’s t-test for measurements of signals evoked by 1 μM CAPS before and after NGF, respectively, for each cell category; *** p$
Figure Legend Snippet: NGF-excitable cells exhibit more robust responses to CAPS. Further analysis was performed on DRG subjected to the protocol described for the red trace in Figure 1 A. Cells were categorised according to whether or not they responded to 20 min. with NGF. ( A ) The number of NGF-excitable (red bars) and -refractory (blue bars) cells that responded to each treatment with the indicated CAPS concentrations applied before [B] or after [A] NGF. Analysis tools were applied to each sub-category to measure ( B ) Lag, ( C ) Duration and ( D ) Max. intensity, in all cases plotted as means ± s.e.m., with blue and red dots representing, respectively, values from individual cells assigned to the NGF-refractory and -excitable categories. Black asterisks represent p values derived by unpaired Student’s t-test between the NGF-excitable and –refractory cells within each CAPS stimulation group. Coloured asterisks show results from paired Student’s t-test for measurements of signals evoked by 1 μM CAPS before and after NGF, respectively, for each cell category; *** p

Techniques Used: Derivative Assay

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1) Product Images from "Ca2+ Signalling Induced by NGF Identifies a Subset of Capsaicin-Excitable Neurons Displaying Enhanced Chemo-Nociception in Dorsal Root Ganglion Explants from Adult pirt-GCaMP3 Mouse"

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