October 1, 1994;
Cardiac troponin I is a heart-specific marker in the Xenopus embryo: expression during abnormal heart morphogenesis.
Cardiac troponin I
) expression is restricted to the heart
at all stages of Xenopus development. Whole-mount in situ hybridization and Northern blot analysis indicates that troponin Ic
is first expressed in tailbud
embryos (stage 28
) about the time of the first cytological heart
differentiation and about 24 hr before beating tissue
is observed. We have used this marker to examine abnormal heart
morphogenesis in embryos treated with retinoic acid and lithium. When retinoic acid is administered to embryos prior to heart
specification, heart tissue
is reduced and often completely ablated. When embryos are treated after heart
specification, but before the heart primordium
migrates to the ventral
midline, the migration is unaffected but smaller, abnormal hearts result. Lithium treatment of cleavage stage
embryos causes an increase in heart tissue
. In severely dorsalized embryos, heart tissue
can be found around the entire embryo
with the exception of a small gap at the most dorsal point. This gap indicates that migration of the heart
to the ventral
midline does not occur in these embryos. Later in development, a centrally located, beating heart
is observed in dorsalized embryos. The timing of its appearance suggests that it is formed by movements normally associated with heart
morphogenesis rather than migration.
[+] show captions
tnni3 <troponin I type 3 (cardiac) > expression in Xenopus laevis embryo, stage 35&36, in situ hybridization, lateral view, anterior left.
Copyright © Paul A. Krieg
FIG. 2. Expression of troponin Ic (blue stain) is limited to the heart in Xenopus embryos. Troponin Ic expression was visualized by whole
mount in situ hybridization using a digoxygenin-labeled antisense troponin Ic RNA. (A) A stage 28-30 Xenopu.5 embryo showing that troponin
Ic is expressed when the heart is still a simple tube. (B) Magnified view of (A) showing that expression is limited to the heart. (C) A stage 36
Xenopu.s embryo showing expression when the heart has the characteristic S shape as a result of looping. (D) Troponin Ic expression in a
differentiated heart (stage 41).
FIG. 3. Expression profile of cardiac troponin I transcripts during
early development. An RNA blot, containing approximately 10 ug of
poly(A)+RNA per lane, was hybridized with random-primed troponin
Ic probes. The positions of the 18S and 28S RNAs are indicated. Lanes
are labeled with numbers corresponding to Xenopus embryonic stages
(Nieuwkoop and Faber, 1967).
FIG. 4. Heart morphology in embryos treated with retinoic acid. The heart morphology of control embryos at stage 38 is seen in (A), with an enlarged view of a control heart in (D). At the equivalent stage, embryos that were treated with 10 I-'M RA at stage 10.5 (B) have hearts which are extremely reduced in size or absent (bottom embryo). An enlarged view (E) shows the significant reduction in heart size in RA-treated embryos. Note that many other anterior structures are also absent. (C) Embryos at the equivalent of stage 38 that were treated with 10uM RA at stage 15. These show almost normal morphology of anterior structures but the hearts are smaller than those in control embryos. The enlarged
view (F) shows a failure of the heart to separate into distinct chambers. In all RA treatments, the heart tissue migrated to the ventral midline.
FIG. 5. Retinoic acid-treated embryos have abnormal hearts. The heart (h) of a normal stage 38 embryo in longitudinal section (A) is multichambered and is flattened against the gut (see Fig. 6). In (B), the heart (h) of the retinoic acid -treated embryo has a single large chamber that does not appear to have any of the normal constrictions or looping associated with normal cardiac morphogenesis. In both embryos anterior is
to the left as indicated by tbe position of the cement gland (cg).
FIG. 6. Retinoic acid effects upon cardiac and lymph heart size. (A)
The location of the cardiac muscle and the lymph heart in a normal
embryo. (B) With increasing doses of retinoic acid, the length of the
cardiac muscle diminishes and the length of the lymph heart increases.
Error bars represent +1 SD.
F IG. 7. Expression of troponin 1c in lithium-treated embryos, (A) The heart in a stage 28/30 control embryo, Note that it has already formed
a simple tube at the ventral midline. (B) A DAI 8 embryo that has an enlarged heart region. Cardiac tissue is absent on the dorsal side of the
embryo and has not fused at the ventral midline. (C) A top view of a DAI 9 embryo showing the distribution of heart tissue (arrowheads).
Although not visible in its entirety in this view, the heart forms a ring that could be detected around the entire circumference with the exception
of a gap (g) on the dorsal side. The heart tissue is just beneath the surface, indicating that it has not involuted. (D) A stage 38 control embryo
with the heart undergoing looping and deformations that result in the final morphology. (E) A DAI 9 embryo viewed from the side. The heart
now appears radial and is moving to the final location at the center of the embryo. The timing of this movement corresponds to the normal
looping and folding events, rather than migration to the ventral midline. (F) A stage 38 DAI 8 embryo with an almost radial heart. The heart
has begun to move to the center of the embryo but the most dorsal side of the embryo (arrow) is still thin.