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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 ubs3Tg ; pkd2tc321/tc321 ; zf350Tg control Long-pec atrioventricular canal IFL
ubs3Tg ; tnnt2atc300b/tc300b ; zf350Tg control Long-pec atrioventricular canal IFL
ubs3Tg ; tnnt2atc300b/tc300b ; zf350Tg chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal IFL
ubs3Tg ; trpv4sa1671/sa1671 ; zf350Tg control Long-pec atrioventricular canal IFL
zf350Tg ; ubs3Tg control Long-pec atrioventricular canal IFL
zf350Tg ; ubs3Tg chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal IFL
zf350Tg ; ubs3Tg + MO1-gata1a control Long-pec atrioventricular canal IFL
zf350Tg ; ubs3Tg + MO1-gata2a control Long-pec atrioventricular canal IFL
zf350Tg ; ubs3Tg + MO1-gata2a chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal IFL
zf350Tg ; ubs3Tg + MO1-tnnt2a control Long-pec atrioventricular canal IFL
zf350Tg ; ubs3Tg + MO1-trpv4 control Long-pec atrioventricular canal IFL
zf350Tg ; ubs3Tg + MO2-cmlc1 control Long-pec atrioventricular canal IFL
zf350Tg ; ubs3Tg + MO2-trpv4 control Long-pec atrioventricular canal IFL
Antibody Labeling Details No data available
Phenotype Details
Fish Conditions Stage Phenotype
ubs3Tg ; pkd2tc321/tc321 ; zf350Tg control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
ubs3Tg ; tnnt2atc300b/tc300b ; zf350Tg control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
ubs3Tg ; tnnt2atc300b/tc300b ; zf350Tg chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal GCaMP expression amount, ameliorated
ubs3Tg ; trpv4sa1671/sa1671 ; zf350Tg control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
zf350Tg ; ubs3Tg chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
zf350Tg ; ubs3Tg + MO1-gata2a control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
zf350Tg ; ubs3Tg + MO1-gata2a chemical treatment: 4alpha-phorbol 12,13-didecanoate Long-pec atrioventricular canal GCaMP expression amount, ameliorated
zf350Tg ; ubs3Tg + MO1-tnnt2a control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
zf350Tg ; ubs3Tg + MO1-trpv4 control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
zf350Tg ; ubs3Tg + MO2-cmlc1 control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
zf350Tg ; ubs3Tg + MO2-trpv4 control Long-pec atrioventricular canal GCaMP expression decreased amount, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Current biology : CB for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Curr. Biol.