Curiosity and a dash of good luck

In May 1980 I took myself and my wife Madeleine to the famous Station Biologique de Roscoff in Brittany. I had been studying epiboly — a process in the gastrulation of some embryos in which an epithelium spreads gradually over the surface of the egg — in the killifish (Fundulus sp.), but thought I really ought to see if epiboly operates in the same way in another teleost.I chose the lumpfish because of its rather large eggs but found it impossible to remove their membranes. The problem was solved by Alain Maron, one of the excellent marins (collectors) at the station, who suggested the blenny (Blennius pholis), a strange little teleost that hangs around the intertidal zone and deposits its eggs on the underside of rocks, from pebbles to enormous boulders. M. Maron showed me some eggs, which contained a translucent embryo with a beautiful pigment pattern, especially in the pectoral fins, reminiscent of Fundulus. Clearly, I had to see the early stages of gastrulation.The next step was to crowd a bunch of blennies into the aquarium in my simple little laboratory (which overlooked a charming old walled garden). I added a pile of flat rocks, fed the blennies chopped fish and waited. With my well-known good luck, by the next morning there were hundreds of eggs on the underside of one of the rocks.Blenny eggs are of a nice size (about 1.2 millimetres in diameter) and sufficiently lucid to reveal the movement of cells. So, I went to work with an ancient dissecting microscope to follow early development. Fortunately, the Blennius egg develops slowly.What struck me soon after gastrulation had begun was something totally unexpected. Two big black pigment spots slowly appeared just lateral to the prospective trunk region of the embryonic axis. I couldn't imagine what they were doing there. The neural crest was surely only just beginning to form. Naturally I was curious; indeed, I was entranced. I postponed the study of epiboly.By then I was late for dinner, so I walked quickly home along the harbor, downed a generous whisky to celebrate my bizarre discovery, had dinner and immediately drove back to the lab. I remember that little trip vividly. There was dense fog, the streets were empty and the Station Biologique was in complete darkness. I opened the gate in the garden wall and groped my way around the ornamental fish pool towards the dark bulk of the laboratory.It was good that I had returned so quickly. The spots had expanded and their distal periphery now had a ragged look, as if they were spreading. So I decided that I'd better stay with them. The spots continued to spread and they were assuming the appearance of a typical spreading epithelial sheet in which the cells now seemed to be arranged in a quasi monolayer. Rifts that slowly widened to form semicircular gaps began to appear between submarginal cells, indicating increasing tension in the sheet. As spreading continued, the gaps widened and some groups of distal peripheral cells began breaking away from the sheet and spreading on their own over the yolk sac away from the sheet and the embryo. This dismembering continued until there were several such directionally moving groups (Figure 1Figure 1).Figure 1A blenny egg showing the pigmented cell mass in the process of breaking up into small cell clusters. Six clusters have already moved away and another is about to do so. The developing embryo can be seen at the top and at the left.View Large Image | View Hi-Res Image | Download PowerPoint SlideSuddenly, it hit me. I had made a discovery! In the excitement I had unconsciously risen and was pacing my little laboratory talking to myself: “Trinkaus, do you realize that you are observing active directional movement of cell clusters in vivo in the embryo for the first time in history?” I wanted to tell somebody but there was no-one to tell. So I told myself. Then, with throbbing heart, I returned to my microscope and resumed my observations. Where would these clusters go?Lacking a camera, I had been making drawings as I watched. Now, however, my drawings began to make scientific sense. I had discovered an important phenomenon and I had better record it well and in detail. I should point out that I was prepared for recognizing the significance of this discovery. John Kolega in our laboratory at Yale was at the time in the midst of his doctoral research on the movement of cell clusters in tissue culture. Nature favors the prepared mind.I spent much of that night following this remarkable phenomenon. The spots, or ‘pigmented cell masses’ as I come to call them, continued to spread slowly, breaking up into a dozen or more cell clusters. The more peripheral ones moved directionally and fairly steadily away from the embryo proper until each side of the yolk sac was dotted with more or less equally spaced black cell clusters. Curiously, movement ceased when the most peripheral ones reached the most ventral region, just short of the mid-ventral line.There is much more to tell. Later, the cells of these scattered epithelial cell clusters transformed into mesenchymal dendritic melanocytes. The clusters disaggregated in a gradient, beginning with those closest to the embryo, and the melanocytes then migrated as individuals toward the pectoral fin bud on each side of the embryo. As they reached the fin bud, they penetrated it and eventually formed therein an intricate and characteristic pigment pattern, the very pattern I had seen when I first observed a Blennius embryo.The following Spring I returned to Roscoff fully equipped optically and photographically and studied with passion all of this and more. I continued to collect eggs in an aquarium but, as the fish did not always cooperate, I had to supplement my supply with frequent trips to the vast intertidal zone (lagreve) of Roscoff in the company of the marins. This was a happy sport, amid the rocks, seaweed and the eggs. Eventually, I published my studies of these remarkable developmental processes in two papers in the Journal of Experimental Zoology.I tell this little story in an effort to convey some of the emotional impact of a simple scientific discovery and the importance of following one's curiosity. But, as is often the case, this discovery depended in large part on a stroke of good luck. If the cells of these spots were not stuffed with black melanin granules they would never have caught my eye.