Hensen’s node

Lakshml treated primitive-streak stage embryos with CN to study the effect on Hensen s node (related to the organization of the embryo). The embryos were incubated for 3 h after treatment the nodes were then excised, washed, and grafted to host chick embryos at the same stage. The results showed an inverse relation between the capacity for Induction of Hensen s node and exposure to CN. 

Hogan, B.L.M., Thaller, C., Eichle, G. 1992. Evidence that Hensen s node is a site of retinoic acid... 

Kintner, C.R., Dodd, J. 1991. Hensen s node induces neural tissue in Xenopus ectoderm. Implications for the action of the organizer in neural induction. Development 113, 1495-1505. 

When the embryo becomes the trilaminar disk, the process of neurulation begins. Most diagrams of this period show the cells of the new middle layer multiplying rapidly from somewhere in or near the anterior end of the primitive streak, diving imder the ectoderm just anterior to it, between the ectodermal and endodermal layers, and spreading anteriolaterally, toward the prochordal plate and to the sides. These cells appear to correspond closely in fxmction to the cells of the amphibian Organizer, the avian Hensen s node, and the zebrafish shield. 

Fig. 6. Stage 5 (HH), the head process has reached its most cranial position and laid down the prospective head territory. The cells of the epiblast overlying the head process form a pseudostratified epithelium, the medullary plate, and the presumptive neural plate. Hensen s node is clearly asymmetrical. The bilateral heart primodia lie lateral and anterior to the node. (See Color Plate)...

Fig. 11. Stage 10 (HH), 10 pairs of somites (36-42h) dorsally anterior neuropore closed, prominent optic vesicles, rhombomere boundary 4/5 formed, and neural folds are closed to almost the level of the node. Ventrally, Hensen s node has regressed almost to the end of the primitive streak (the 10th pair of somites has not fully segmented caudally in this illustration), pronephric tubules develop between somites 6 and 10, heart tube turns asymmetrical bulging out to the right and contractions can be seen, and bilateral vitelline veins fan out toward the area opaca, which shows large blood islands to establish circulation.

In the following sections, I consider two examples of operations on Hensen s node excision from a donor quail embryo and transplantation to the inner ring of the area opaca of a host chick embryo to demonstrate embryonic induction, as done by Storey et al. (10), and rotation of the node about its rostrocaudal axis in situ, to demonstrate embryonic regulation as done by Abercrombie (14). 

Demonstration of Embryonic Induction by Transplantation of Hensen s Node... 

Fig. 1. Preparation of host and donor embryos for a Hensen s node graft in New culture. A Materials. B-F. Clean the yolk and place it into the dish with saline. G-I. Cut the vitelUne membrane around the yolk.

Fig. 3. Manipulations under the microscope. A Host embryo (stage 3+). B-D Clear yolky endoderm to expose the epiblast at the future grafting site. E Donor embryo, ventral side uppermost (stage 4). F-J Cut out Hensen s node from the donor. K and L Pick up the excised node with a Gilson pipet. M-N Manipulate the node to the graft site. O The finished operation.

The procedure outlined here is generic , and similar manipulations can be done to operate on smaller or larger portions of embryo. For example, whole, large sections of the primitive streak may be rotated as done by Abercrombie (14) and others or very small subsectors of Hensen s node transplanted as described by Selleck and Stern (19) and Storey et al. (9). 

Still observing under the microscope, carefully and slowly withdraw as much saline as possible from inside and outside the ring, as described for grafting Hensen s nodes. In this experiment, where the manipulated node is not secured under a flap of tissue, it is much easier to lose it while sucking off the fluid. If necessary, replace it in position as required, using the fine needles. 

Operated embryos may be analyzed by histology, whole-mount immuno-histochemistry, and in-situ hybridization with appropriate probes, as described for Hensen s node grafts. 

Selleck MAJ, Stem CD (1991) Eate mapping and cell lineage analysis of Hensen s node in the chick embryo. Development 112 615-626. 

Storey KG, Selleck MAJ, Stem CD (1995) Induction by different subpopnlations of cells in Hensen s node. Development 121 417 28. 

Streit A, Stem CD, Thery C, Ireland GW, Aparicio S, Sharpe M, Gherardi E (1995) A role for HGF/SF in neural induction and its expression in Hensen s node during gastrulation. Development 121 813-824. 

SeUeek MAJ, Stem CD (1992) Commitment of mesoderm cells in Hensen s node of the chick embryo to notochord and somite. Development 114 403 15. 

As described for operations on Hensen s node, it is important to avoid damaging the vitelline membrane at all costs. Any leakage of the albumen culture fluid to the inside of the ring diminishes survival and may cause the grafted tissue to fall out of its site. 

Catala, M., Teillet, M.-A., De Robertis, E. M., and Le Douarin, N. M. (1996) A spinal cord fate map in the avian embryo while regressing, Hensen s node lays down the notochord and floor plate thus joining the spinal cord lateral walls. 

Chen, Y., Huang, L., Russo, A. E, and Solursh, M. (1992) Retinoic acid is enriched in Hensen s node and is developmentally regulated in the early chicken embryo. Proc. Natl. Acad. Sci. USA 89,10,056-10,059. 

Eirst Insert Eigure 2, chapter 18, Grafting Hensen s Node, by Claudio D. Stem. 

---