ZFIN ID: ZDB-FIG-160726-4
Heckel et al., 2015 - Oscillatory Flow Modulates Mechanosensitive klf2a Expression through trpv4 and trpp2 during Heart Valve Development. Current biology : CB   25(10):1354-1361 Full text @ Curr. Biol.
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Gene:
Fish:
Condition:
Knockdown Reagents:
Anatomical Term:
Stage: Long-pec
PHENOTYPE:
Fish:
Condition:
Knockdown Reagents:
Observed In:
Stage: Long-pec

Fig. 4

Endocardial Intracellular Calcium Level Correlates with the Intracardiac FH Index and Depends on TRPs Channels Function

(A and B) Calcium level quantification in the embryonic heart of controls (n = 10), mutants of heart contraction (cmlc1 knockdown; n = 3), lidocaine-treated embryos (n = 3), gata1 (n = 3), gata2 (n = 6) MO, and gata2 MO plus 4α-PDD (n = 2). At 48 hpf, fishes with a reduced oscillatory flow index (sih, lidocaine, cmlc1, and gata2) show a significant reduction of intracellular calcium intensity in the AVC (white arrow), whereas the specific ligand of TRPV4 (4α-PDD) channels restores an almost normal intracellular calcium intensity in the AVC of the gata2 MO. *p < 0.05; **p < 0.01 ANOVA. Error bars indicate the SD.

(C) Calcium level quantification in the embryonic heart of control, trpp2 MO (n = 3), trpp2cup-/- (n = 6), trpv4 MO (n = 6), and trpv4sa1671-/- (n = 2) embryos. Error bars indicate the SD.

(D) 4α-PDD induces an increase in intracellular calcium in the absence of flow in the Tg(fli1a:gcamp3.0), sih (n = 5) hearts. ***p < 0.001 t test. Error bars indicate the SD.

(E) Confocal micrograph showing the effect of 4α-PDD on calcium signaling in sih.

(F) Confocal micrograph and intensity quantification of klf2a:H2B-eGFP intensity in gata2 knockdown with and without 4α-PDD treatment (n = 9 and n = 3, respectively). Error bars indicate the SD.

(G) Schematic highlighting a potential mechanism for oscillatory flow mechanodetection by the TRP channels (in red) specifically in the AVC.

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
GCaMP pkd2tc321/tc321; ubs3Tg; zf350Tg control Long-pec atrioventricular canal IFL
tnnt2atc300b/tc300b; ubs3Tg; zf350Tg control Long-pec atrioventricular canal IFL
tnnt2atc300b/tc300b; ubs3Tg; zf350Tg chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal IFL
trpv4sa1671/sa1671; ubs3Tg; zf350Tg control Long-pec atrioventricular canal IFL
ubs3Tg; zf350Tg control Long-pec atrioventricular canal IFL
ubs3Tg; zf350Tg chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal IFL
ubs3Tg; zf350Tg + MO1-gata1a control Long-pec atrioventricular canal IFL
ubs3Tg; zf350Tg + MO1-gata2a control Long-pec atrioventricular canal IFL
ubs3Tg; zf350Tg + MO1-gata2a chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal IFL
ubs3Tg; zf350Tg + MO1-tnnt2a control Long-pec atrioventricular canal IFL
ubs3Tg; zf350Tg + MO1-trpv4 control Long-pec atrioventricular canal IFL
ubs3Tg; zf350Tg + MO2-cmlc1 control Long-pec atrioventricular canal IFL
ubs3Tg; zf350Tg + MO2-trpv4 control Long-pec atrioventricular canal IFL
Antibody Labeling Details No data available
Phenotype Details
Fish Conditions Stage Phenotype
pkd2tc321/tc321; ubs3Tg; zf350Tg control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
tnnt2atc300b/tc300b; ubs3Tg; zf350Tg control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
tnnt2atc300b/tc300b; ubs3Tg; zf350Tg chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal GCaMP expression amount, ameliorated
trpv4sa1671/sa1671; ubs3Tg; zf350Tg control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
ubs3Tg; zf350Tg chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
ubs3Tg; zf350Tg + MO1-gata2a control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
ubs3Tg; zf350Tg + MO1-gata2a chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal GCaMP expression amount, ameliorated
ubs3Tg; zf350Tg + MO1-tnnt2a control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
ubs3Tg; zf350Tg + MO1-trpv4 control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
ubs3Tg; zf350Tg + MO2-cmlc1 control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
ubs3Tg; zf350Tg + MO2-trpv4 control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
Acknowledgments:
This image is the copyrighted work of the attributed author or publisher, and ZFIN has permission only to display this image to its users. Additional permissions should be obtained from the applicable author or publisher of the image. Full text @ Curr. Biol.