The postembryonic growth of the compound eye of the cockroach Periplaneta americana involves increases in the size of the individual ommatidia as well as a 35-fold increase in the number of ommatidia. These ommatidia are added to the anterior, dorsal, and ventral margins of the eye by means of an almost continuous process of cell division in the proliferation zone in these margins. This proliferation phase is followed by a process of maturation of bundles of ‘pre-ommatidial’ cells into mature ommatidia, a process which involves further cell division. Processes involved in compound-eye development are investigated by eye margin grafting, histological techniques and cell proliferation studies.

Following the emergence of a hemimetabolous insect from the egg, the compound eye grows from stadium to stadium by a combination of increases in ommatidial cell size and increases in total numbers of ommatidia (Bodenstein, 1953). Some insects such as Dixippus morosus add very few facets to their growing eye (Friza, 1928) whereas others such as Sphodromantis bioculata (Yamanouti, 1933) and the dragonflies (Ando, 1957; Sherk, 1977, 1978a, b) exhibit a spectacular increase in ommatidial numbers during their development from nymph to adult. The compound eye of a newly hatched Periplaneta americana nymph contains just over 100 ommatidia, while the adult eye can contain over 3500 (Fig. 1).

Fig. 1

The right compound eye (ce) of a P. americana adult. The inset shows a newly hatched larva photographed at the same magnification. The larval eye contains just over 100 ommatidia while that of the adult contains over 3500. Growth of the compound eye occurs both by increase in size of the ommatidia and by addition of new ommatidia. Specimens were prepared by freeze drying to show individual facets clearly, a, antenna; o, lateral ocellus. Bar represents 0·25 mm.

Fig. 1

The right compound eye (ce) of a P. americana adult. The inset shows a newly hatched larva photographed at the same magnification. The larval eye contains just over 100 ommatidia while that of the adult contains over 3500. Growth of the compound eye occurs both by increase in size of the ommatidia and by addition of new ommatidia. Specimens were prepared by freeze drying to show individual facets clearly, a, antenna; o, lateral ocellus. Bar represents 0·25 mm.

We have shown (Nowel & Shelton, 1980) that the eye margin of the cockroach does not advance through adjacent head epidermis to recruit it into the expanding compound eye. Rather, it acts as a ‘budding zone’ which generates cells to form new ommatidia. In the present study, the histology of the edge of the growing eye plus the levels of cell division in the eye and head epidermis are examined at various points of the moult cycle. The results of these observations further substantiate the role of the eye margin as a budding zone.

Hyde (1972) has studied the appearance of the developing compound eyes of various stadia in the cockroach P. americana and has concluded that growth occurs all around the perimeter of each eye, using as her criterion the presence of an unpigmented zone seen to surround the eyes of living animals (Fig. 2, 3 c, d). Maturation of ommatidia and their addition to pre-existing ones already present in the previous instar was said to occur in this zone of maturation which surrounds the eye. Anderson (1976) has since offered histological evidence to contradict this: histological sections show that zones of growth and maturation are not present along the posterior margin of the eye, but are confined to the anterior and dorsal borders. The experiments to be described were designed to distinguish between the two alternatives. The results of these experiments are in agreement with Anderson’s (1976) histological observations: the compound eye of the cockroach grows along its dorsal, anterior, and ventral faces, but not along its posterior edge.

Figure 2

A chimeric compound eye of P. americana generated by implanting a graft of wild-type eye margin into the eye of a la vender nymph. The chimera was photographed at each post-operative stadium up to and including the adult (3 g). The series shows the locations of the older and younger ommatidia and the numbers of rows of ommatidia added to the eye during each stadium. Note that more growth occurs in the anterior than in the posterior eye margin. Note also the stability of the numbers of ommatidia along the posteroventral graft/host border (14 ommatidia at each stadium) and the increase in ommatidia along the anteroventral border during the post-opeiative stadia. This demonstrates that no new ommatidia are generated by the posterior eye margin whereas the dorsal and anterior (as well as the ventral) eye margins produce new ommatidia during posternbryonic development. A, anterior; D, dorsal; g, graft-derived ommatidia; h, host-derived ommatidia; o, lateral ocellus. Bar represents 0·25 mm.

Figure 2

A chimeric compound eye of P. americana generated by implanting a graft of wild-type eye margin into the eye of a la vender nymph. The chimera was photographed at each post-operative stadium up to and including the adult (3 g). The series shows the locations of the older and younger ommatidia and the numbers of rows of ommatidia added to the eye during each stadium. Note that more growth occurs in the anterior than in the posterior eye margin. Note also the stability of the numbers of ommatidia along the posteroventral graft/host border (14 ommatidia at each stadium) and the increase in ommatidia along the anteroventral border during the post-opeiative stadia. This demonstrates that no new ommatidia are generated by the posterior eye margin whereas the dorsal and anterior (as well as the ventral) eye margins produce new ommatidia during posternbryonic development. A, anterior; D, dorsal; g, graft-derived ommatidia; h, host-derived ommatidia; o, lateral ocellus. Bar represents 0·25 mm.

Figure 3

Chimeric compound eyes of P. americana showing the growth at the eye margins. By comparing the numbers of ommatidia between a fixed position (the graft/host border) and the eye margin in the adults (b, d) with those in young larvae (a, c), it is apparent that new ommatidia have been added at the anterior, dorsal and anterodorsal eye margin. Counts of numbers of facets between a particular ommatidium (arrows, Figs. 3 c, d) and the posterior margin of the eye always remained constant from moult to moult. No new ommatidia are produced at the posterior margin. Note the band of unpigmented tissue surrounding the eye, which Hyde (1972) called the ‘growing zone’. Note also the asymmetry of eye growth: more growth occurs in the anterior than in the posterior regions of the eye, as determined by the longer graft/host border at the anterior edge of the graft.

(a) Larva after a single post-operative moult.

(b) Same animal photographed in (a) but after six post-operative moults to the adult.

(c) Larva after two post-operative moults.

(d) Same animal photographed in (c) but after six post-operative moults to the adult.

A, anterior; D, dorsal; g, graft-derived ommatidia; h, host-derived ommaditia. Bar represents 0·25 mm.

Figure 3

Chimeric compound eyes of P. americana showing the growth at the eye margins. By comparing the numbers of ommatidia between a fixed position (the graft/host border) and the eye margin in the adults (b, d) with those in young larvae (a, c), it is apparent that new ommatidia have been added at the anterior, dorsal and anterodorsal eye margin. Counts of numbers of facets between a particular ommatidium (arrows, Figs. 3 c, d) and the posterior margin of the eye always remained constant from moult to moult. No new ommatidia are produced at the posterior margin. Note the band of unpigmented tissue surrounding the eye, which Hyde (1972) called the ‘growing zone’. Note also the asymmetry of eye growth: more growth occurs in the anterior than in the posterior regions of the eye, as determined by the longer graft/host border at the anterior edge of the graft.

(a) Larva after a single post-operative moult.

(b) Same animal photographed in (a) but after six post-operative moults to the adult.

(c) Larva after two post-operative moults.

(d) Same animal photographed in (c) but after six post-operative moults to the adult.

A, anterior; D, dorsal; g, graft-derived ommatidia; h, host-derived ommaditia. Bar represents 0·25 mm.

Maintenance of cockroach stocks

Cultures of P. americana were maintained under conditions of constant temperature (24°C) and an alternating cycle of 12 h light/12 h dark, and fed on a diet of rat pellets and water.

Surgical techniques

Newly moulted third-fifth instar nymphs were selected for operations, anaesthetized in small glass vials cooled on ice for 10–20 min, and immobilized on a bed of plasticine. Excisions of integument grafts were made using a razor blade fragment (Gillette français) supported in a pin vice, and transferred to homotopic sites prepared in host animals by removing integument of equal size and shape. The grafts, consisting of eye margin and adjacent head epidermis, were held in place using a small droplet of melted insect wax (Krogh & WeisFogh, 1951). Wild-type and lavender (Ross, Cochran & Smyth, 1964) stocks of P. americana were used in graft exchanges.

Tissue preparation

Eye material was prepared by fixing in alcoholic Bouin, embedded in paraffin, sectioned at 10 μm and stained with Delafield’s haematoxylin and eosin (Pantin, 1969). Alternatively, material was fixed in a glutaraldehyde/paraformaldehyde mixture (Karnovsky, 1965) buffered in a phosphate buffer (Hayat, 1970) at pH 7·4 for 2–4 h, and then postfixed in phosphate-buffered 1% osmium tetroxide for 2–3 h, following which it was dehydrated in an acetone series and embedded in Spurr’s resin. Semithin (1 pm) sections were cut using a Huxley Ultramicrotome with glass knives, and stained with 1% toluidine blue in 1% borax.

Colchicine studies

In order to investigate the temporal aspects of cell proliferation within the developing eye during the moult cycle, and to locate areas where cell division occurs, a 1% colchicine solution (0·5 μl per 0·1 g live weight of animal) made up in insect saline (Hoyle, 1953) was injected into cockroach nymphs through a drawn micropipette inserted into an antenna. P. americana larvae of inter-mediate stages (fifth, sixth & seventh instar nymphs) were injected at different points during their moult cycle (0, 1, 3, 5, 7, 9, 14, 21 and 28 days following the previous moult). Nymphs at these intermediate stages have an intermoult period approximately one month in duration (Biellmann, 1960), and so consideration of the 28-day animals was restricted to those nymphs which were about to moult as determined by a cloudy appearance of the eyes (Flint & Patton, 1959). Following injection with colchicine, the cut antenna was sealed with insect wax (Krogh & Weis-Fogh, 1951); 12 h later, the animals were fixed for wax histology (three-six animals for each of the nine stages). Paraffin-embedded specimens were cut in a plane perpendicular to the growing dorsal edge of the eye. For a quantitative investigation, every third 10μm section was examined for mitotic figures.

Dividing cells were scored in each of four areas: the head epidermis adjacent to the eye margin (a zone of epidermis extending 375 μm from the border of the eye -an arbitrary but convenient distance for examining microscopically); the proliferation zone of the eye margin (P.Z.); the maturing zone of the eye (M.Z.) the mature eye. The appearance and location (at the cuticular inner surface) of dividing cells is the same in all four regions examined. (Additional samples, some treated with colchicine and some untreated, from each stage were fixed and embedded in Spurr’s resin and sectioned at 1 μm for light microscopy.)

Photography

Experimental animals were photographed on a Zeiss Tessovar Photomacrographic Zoom system. Sectioned material was photographed on a Zeiss Photomicroscope II.

Location of the growth zone in the eyes of P. Americana

Of the 50 nymphs on which operations had been performed to exchange wild-type with lavender eye margins, 39 survived to the imago. Four of these showed no donor-pigmented ommatidia and were discarded. Thirty-five animals had a patch of ommatidia with donor-specific pigment in their compound eyes. This graft-derived eye tissue was visible from the first post-operative moult. As the eye grew along its margins, donor-phenotype ommatidia were added by the implanted segment of eye margin, while host ommatidia were added by the native eye margin. A clear boundary between graft- and host-derived tissues, visible on the basis of pigment differences, is thus formed between eye tissues of these different origins. These boundary lines effectively demonstrate the directions of eye expansion (Figs. 2, 3). In order to visualize this expansion, the graft borders of these 35 mature chimeric eyes were projected onto a photograph of an adult eye, and traced onto the corresponding region of the photograph (Fig. 4).

Fig. 4

Photograph (a) and drawing (b) show the dorsal half of the right compound eye of P. americana. Outlines of graft/host borders of a number of adult chimeras generated by homotopic grafting between wild-type and lavender nymphs have been traced onto these pictures to demonstrate the direction of postembryonic growth of the compound eye. The growth of this half of the eye is predominantly in an anterodorsal direction. A, anterior; D, dorsal. Bar represents 0·5 mm.

Fig. 4

Photograph (a) and drawing (b) show the dorsal half of the right compound eye of P. americana. Outlines of graft/host borders of a number of adult chimeras generated by homotopic grafting between wild-type and lavender nymphs have been traced onto these pictures to demonstrate the direction of postembryonic growth of the compound eye. The growth of this half of the eye is predominantly in an anterodorsal direction. A, anterior; D, dorsal. Bar represents 0·5 mm.

In the dorsal half of the eyes examined in this series of experiments, growth is most extensive in the anterodorsal region of the compound eye. Here there is the greatest increase in the linear dimension of the margin resulting in the divergence of graft/host border lines as they approach the margin (see Fig. 4), and the numbers of rows of new ommatidia added radially in each succeeding stadium is at its greatest in the anterodorsal quadrant. Relatively few rows of new ommatidia are added to the posterodorsal margin per moult (see Figs. 2, 3).

Of the 35 experimental animals, the chimeric compound eyes of 10 were photographed following each post-operative moult. From analysis of the complete series of photographs, it is possible to define the extent of the growing margin and the patterns of growth in different portions of the eye. Numbers of ommatidia added per moult in different parts of the eye were noted, and it was possible to differentiate the growth resulting from addition of new ommatidia from that resulting from enlargement of pre-existing ones.

Because particular ommatidia are recognizable by their position with respect to the stable graft/host border, their position with respect to the changing eye/ head-epidermis border can be followed. In succeeding stadia, particular facets become further and further separated from the anterior, dorsal, anterodorsal and ventral eye margins by more and more newly added rows of facets. This was noted in each of the ten chimerae produced. Ommatidial counts from the graft/host border to these margins demonstrate addition of ommatidia during the post-operative instars. (Owing to the curvature of the ventral portion of the eye, however, photographic presentation of these data is difficult.) No new ommatidia are added to the posterior margin of the eye (Figs. 2, 3). These results support the findings of Anderson (1976) against those of Hyde (1972).

Histology of the growing eye margin

Examination of sections through the edge of the eyes of P. americana nymphs shows retinal elements in various stages of developmental organization (Fig. 5). The proliferation zone is located at the extreme border of the eye along its dorsal, anterior and ventral faces. Its cells are recognised by their undifferentiated and ungrouped appearance and by their dense packing. Mature ommatidia have all their component cells in their correct proportions and positions, and they are found in the central and posterior part of the retina. The region between the proliferation zone and the zone of mature ommatidia shows preommatidia in various stages of differentiation. This zone of maturing ommatidia is operationally defined as a band of preommatidial bundles/ ommatidia six bundles wide (approximately six rows of ommatidia are added to the growing retina per moult during these intermediate larval instars: see Fig. 2).

Figure 5

Micrographs showing semithin sections through the left eye and head epidermis of P. americana nymphs fixed at different times during the intermoult period show the changing eye/epidermis interface.

(a) Newly moulted. The proliferation zone (PZ) and maturation zone (MZ) are visible in the eye. The compound eye is not clearly separated from the epidermis (E) at this stage, c, cuticle.

(b) 7 days post ecdysis. The basement membrane (bm) of the eye is visible between the eye margin and a fold of epidermis adjacent to it. The eye tissue extends to a small cuticular ridge (arrow) which marks the boundary between it and the epidermis. c, cuticle; M.Z., maturation zone; P.Z. proliferation zone.

(c) 28 days post ecdysis. The cuticular ridge (arrow) is extended and the proliferation zone (P.Z.) underlies it. Note the epidermal bristle (b) penetrating the cuticle (c) at the eye margin. Its appearance indicates that the head epidermis is being pushed ahead rather than being recruited by the expanding eye. M.Z. maturation zone.

(d) Pre-apolysis. Note the continued clear division between the eye and head epidermis (E). The folded appearance of the epidermis lying ahead of the cuticular ridge (arrow) suggests it is being pushed back rather than being recruited by the growing eye. c, cuticle; M.Z. maturation zone; P.Z. proliferation zone.

(e) At apolysis, which marks the beginning of the moulting process, new cuticle (nc) deposition has just begun. Note the extent of the cuticular ridge (arrow) of the old cuticle (pc). E. epidermis; M.Z., maturation zone; P.Z. proliferation zone.

(f) Later as the moult is approached, the inner (new) cuticle (nc) is thickened. The old, shedding cuticle has been lost in tissue preparation. E, epidermis; M.Z. maturation zone; P.Z., proliferation zone.

Bars represent 10μm.

Figure 5

Micrographs showing semithin sections through the left eye and head epidermis of P. americana nymphs fixed at different times during the intermoult period show the changing eye/epidermis interface.

(a) Newly moulted. The proliferation zone (PZ) and maturation zone (MZ) are visible in the eye. The compound eye is not clearly separated from the epidermis (E) at this stage, c, cuticle.

(b) 7 days post ecdysis. The basement membrane (bm) of the eye is visible between the eye margin and a fold of epidermis adjacent to it. The eye tissue extends to a small cuticular ridge (arrow) which marks the boundary between it and the epidermis. c, cuticle; M.Z., maturation zone; P.Z. proliferation zone.

(c) 28 days post ecdysis. The cuticular ridge (arrow) is extended and the proliferation zone (P.Z.) underlies it. Note the epidermal bristle (b) penetrating the cuticle (c) at the eye margin. Its appearance indicates that the head epidermis is being pushed ahead rather than being recruited by the expanding eye. M.Z. maturation zone.

(d) Pre-apolysis. Note the continued clear division between the eye and head epidermis (E). The folded appearance of the epidermis lying ahead of the cuticular ridge (arrow) suggests it is being pushed back rather than being recruited by the growing eye. c, cuticle; M.Z. maturation zone; P.Z. proliferation zone.

(e) At apolysis, which marks the beginning of the moulting process, new cuticle (nc) deposition has just begun. Note the extent of the cuticular ridge (arrow) of the old cuticle (pc). E. epidermis; M.Z., maturation zone; P.Z. proliferation zone.

(f) Later as the moult is approached, the inner (new) cuticle (nc) is thickened. The old, shedding cuticle has been lost in tissue preparation. E, epidermis; M.Z. maturation zone; P.Z., proliferation zone.

Bars represent 10μm.

Examining sections through eyes at various points during the moult cycle shows the development of a cuticular ridge under which the growing eye margin advances. The epidermis adjacent to the eye margin (see Fig. 56-d) has the appearance of being displaced by the expanding eye and the extending ridge. The development of the ridge (the extent of which is best seen at ecdysis: Fig. 5e) shows that the eye remains distinct from the epidermis even as the eye expands during the intermoult period.

Location and identification of mitotic figures

Typical sections through the eyes of colchicine-treated P. americana are shown in Fig. 6. Graphs showing the levels of mitotic activity in various regions of the eye and head epidermis through the moult cycle following colchicine treatment are shown in Fig. 7. Cells in the four regions of investigation (proliferation zone, maturation zone, mature eye and head-capsule epidermis) which are undergoing mitosis are recognizable in histological sections according to the following criteria: (a) their basal ends are detached from the basement membrane; (b) the cells are rounded up just beneath the cuticle, and (c) they exhibit a basophilic condensation of chromosome material.

Figure 6

(a) Micrograph of a semithin horizontal section through the eye margin of a wild type P. americana nymph to show the location of dividing cells (arrows). Nine days after moulting, the animal was treated with colchicine to arrest cell division, and fixed 12 h later. Mitotic figures are seen in the proliferation zone (P.Z.) and in the maturation zone (M.Z.). Note the cuticular ridge (cr) separating the head epidermis from the eye. (6) Micrograph of a 10 μm thick section through the right eye and head epidermis of a P. americana nymph treated with colchicine in the middle of the intermoult period (14 days post ecdysis). This period falls within the plateau period of high mitotic activity for the proliferation zone (P.Z.) and in the peak periods of mitotic activity for the zone of maturing ommatidia (M.Z.), the mature retina (M.R.) and the head capsule epidermis (E). Mitotic figures (arrows) are visible in all these areas. Note also the elongated cuticular ridge (cr) along which the preommatidia develop. It separates the retina from the head epidermis.

Bars represent 10μm.

Figure 6

(a) Micrograph of a semithin horizontal section through the eye margin of a wild type P. americana nymph to show the location of dividing cells (arrows). Nine days after moulting, the animal was treated with colchicine to arrest cell division, and fixed 12 h later. Mitotic figures are seen in the proliferation zone (P.Z.) and in the maturation zone (M.Z.). Note the cuticular ridge (cr) separating the head epidermis from the eye. (6) Micrograph of a 10 μm thick section through the right eye and head epidermis of a P. americana nymph treated with colchicine in the middle of the intermoult period (14 days post ecdysis). This period falls within the plateau period of high mitotic activity for the proliferation zone (P.Z.) and in the peak periods of mitotic activity for the zone of maturing ommatidia (M.Z.), the mature retina (M.R.) and the head capsule epidermis (E). Mitotic figures (arrows) are visible in all these areas. Note also the elongated cuticular ridge (cr) along which the preommatidia develop. It separates the retina from the head epidermis.

Bars represent 10μm.

Fig. 7

Bar graphs to show the mitotic activity in (A) the proliferation zone and (B) the maturation zone of the eye margin, and (C) the head epidermis adjacent to the retina of P. americana nymph. Numbers of mitotic figures per examined section were plotted against the number of days after the previous moult on which the animals were injected with colchicine. Each bar represents the average number of mitotic figures/section derived from the examination of at least 100 sections from at least four different animals.

(A) In the proliferation zone, dividing cells are observed on all examined days of the moult cycle. From a low point just before ecdysis, numbers of divisions steeply rise to a plateau from days 3–14, and then gradually fall again.

(B) Dividing cells in the region of maturing ommatidia appear during a restricted portion of the moult cycle only. Cell divisions begin when one quarter of the intermoult period has passed, and end when the animal is about to moult. This pattern is similar to that qualitatively observed (but not quantified for technical reasons) in the mature retina.

(C) Dividing cells in the head epidermis are seen during a similarly restricted portion of the moult cycle. The pattern of cell divisions in the head epidermis (as well as in the regions of maturing and mature ommatidia) is therefore different from that in the proliferation zone of the eye margin.

Fig. 7

Bar graphs to show the mitotic activity in (A) the proliferation zone and (B) the maturation zone of the eye margin, and (C) the head epidermis adjacent to the retina of P. americana nymph. Numbers of mitotic figures per examined section were plotted against the number of days after the previous moult on which the animals were injected with colchicine. Each bar represents the average number of mitotic figures/section derived from the examination of at least 100 sections from at least four different animals.

(A) In the proliferation zone, dividing cells are observed on all examined days of the moult cycle. From a low point just before ecdysis, numbers of divisions steeply rise to a plateau from days 3–14, and then gradually fall again.

(B) Dividing cells in the region of maturing ommatidia appear during a restricted portion of the moult cycle only. Cell divisions begin when one quarter of the intermoult period has passed, and end when the animal is about to moult. This pattern is similar to that qualitatively observed (but not quantified for technical reasons) in the mature retina.

(C) Dividing cells in the head epidermis are seen during a similarly restricted portion of the moult cycle. The pattern of cell divisions in the head epidermis (as well as in the regions of maturing and mature ommatidia) is therefore different from that in the proliferation zone of the eye margin.

Cell division in the proliferation zone

The P.Z. in the eye margin is the only one of the four areas studied showing mitoses on all of the days examined (Fig. 7 a). The level of mitotic activity in this region reaches a maximum three days after the proceeding moult, rising sharply from day 0. This high level is maintained over a period of approximately two weeks (analysis of the results showed that the levels of mitosis are similar on days 3, 5, 7, 9, and 14). Following this plateau phase, the level gradually falls to a low point towards the end of the stadium.

At one point in the moult cycle the level of cell division within the P.Z. (and also in the M.Z., the mature retina, and the head epidermis) falls to zero. This point occurs shortly before moulting. The precise chronological point has not been defined because individual animals develop at slightly different rates1.

In 28-day animals which are about to moult as shown by a thick (35–40 μm) newly deposited cuticle underlying the old cuticle which is about to be sloughed off, cell divisions are seen in the P.Z. The same age animals with a slightly slower rate of development (as shown by a thinner (5–10 /on) newly deposited cuticle) show no cell divisions in structures derived from head ectoderm. It is clear that cell division ceases at the start of new cuticle deposition. Shortly thereafter, and even before ecdysis has occurred, cell division in the proliferation zone resumes.

Cell division in the maturing zone, the area of mature ommatidia, and the head epidermis

In these three regions, 80% of the mitotic figures were found during a limited part of the moult cycle, namely at 9 days and 14 days after the previous moult or during the second quarter of the intermoult period (Fig. lb, c). However, one 21-day animal showed an abnormal level of cell division compared with the three others examined in that group which showed virtually no mitoses. Considering the variability of the actual length of the moult cycle1, such a 21-day animal could be one developing at a slower rate which is still in the first half of its intermoult period. If this individual were to be eliminated from consideration, then the mitoses occurring at days 9 and 14 into the (presumably 28-day) intermoult period would comprise 97% of the total mitoses in these regions for the complete moult cycle.

The dimensions of the compound eye increase from instar to instar by addition of new ommatidia along its dorsal, anterior, and ventral borders. Particular (marked) ommatidia become further and further removed from these margins of the eye while having the same number of facets separating them from the posterior margin. Growth in this direction is due to increase in ommatidial size only.

Hyde’s (1972) conclusion that the zones of growth and maturation completely surround the retina must therefore be incorrect. The zone of growth lies only along the dorsal, anterior and ventral borders of the compound eye (Anderson, 1978).

A significant observation from these studies is that different regions of the growth zone produce ommatidia at different rates. There is more growth, for example, along the anterodorsal edge of the eye than along the posterodorsal edge (see Figs. 2, 3; for instance, the specimen illustrated in Fig. 2 showed that, during the post-operative instars to the adult, 49 rows of ommatidia were added along the graft/host border to the anterodorsal edge of the eye, while only 29 rows were added to the posterodorsal edge). This suggests that regions along the eye margin have some inherent properties or are under some regional control to govern the rate of production of new ommatidia and therefore the rate of expansion of the eye.

The pattern of mitoses in the P.Z. of the eye margin is different from that of the M.Z., the mature ommatidia, and the head epidermis (compare Fig. 7a-c). During the entire intermoult period, the cells of the P.Z. undergo only a brief cessation of division, immediately before moulting. In contrast, the cells of the other three regions considered, more closely resemble the pattern found in a wide variety of other ectodermal epithelia (Rhodnius prolixus abdominal epithelium: Locke, 1964; Schistocercagregaria abdominal epithelium: Anderson, 1976, 1978; Leucophaea maderae abdominal epithelium: Bray 1978). Here cell division is confined to a discrete period within the moult cycle. Following ecdysis (or in the case of R. proxilus, the blood meal) there is no cell division for the first one fifth to one third of the intermoult period. After this interval, mitotic activity begins and continues until the intermoult period is approximately half over. At this point, the level of mitosis falls to zero where it stays for the remainder of the moult cycle.

The results for the head epidermis of P. americana described here are comparable to those described above, and mitoses in the M.Z. and mature ommatidia follows a similar pattern. It seems reasonable to assume that cell divisions in these three regions are under the same (possibly hormonal) control, though the precise nature of these controls remains obscure.

The fundamentally different pattern of mitotic activity in the P.Z. (which extends around the anterior, dorsal, and ventral borders of the eye) provides further evidence that it is a specialized source of new cells which are required throughout the moult cycle for eye growth, i.e. a ‘budding zone’ for the production of cells to form new ommatidia in the expanding eye (Bodenstein, 1953).

During its postembryonic development, a continuous sequence of events occurs in the growing zone of the retina to transform undifferentiated cells into mature ommatidia. In the locust S. gregaria, five stages of development can be identified (Eley & Shelton, 1976): (a) Ungrouped cells. These are seen as a band of closely packed, undifferentiated cells immediately adjacent to the eye/ head epidermis border. (b) Early cell clusters. Cells are grouped into rows of bundles, or ‘pre-ommatidia’ (Imberski, 1967). (c) Late cell clusters. Cytological differentiation has made component cells in the pre-ommatidia distinguishable. (d) The developing rhabdom stage. Retinula cells form a rosette with microvilli from each cell emanating centrally to form the rhabdom. (e) The mature ommatidium. This consists of four distal cone cells with narrow cone-cell processes, two primary pigment cells and a variable number (7-13) of secondary pigment cells, and eight retinula cells surrounding a mature rhabdom (Eley & Shelton, 1976; Eley, 1978).

The formation of this final pattern in S. gregaria involves a programme of mitosis in the eye similar to that found in the present studies on P. americana. That is, the proliferation zone is characterized by a high level of mitotic activity throughout the intermoult period. Virtually all mitotic activity in the zone of ommatidial maturation occurs during the time mitoses in the P.Z. are at their peak, as do the mitoses in the mature retina (Anderson, 1978).

In Drosophila, the undifferentiated cells of the eye imaginai disc are transformed into ommatidia following two waves of mitoses. The first mitotic wave results in “pre-clusters” of cells (retinula cells, 3, 3, 4, 5, 8) becoming post-mitotic. The second wave of mitoses results in the formation of the remaining cells required to complete the ommatidium, i.e., cone cells, pigment cells, bristle cells plus retinula cells 1, 6, 7 (Ready, Hanson & Benzer, 1976).

In P. americana there appears to be a single wave of mitoses (in the P.Z.) followed by a pulse of mitotic activity (in the M.Z.) which occurs once during each moult cycle. In contrast to the travelling waves of mitoses in Drosophila, which are described as passing across the eye disc (Ready et al. 1976), the wave of cell division in the cockroach is a standing wave, remaining at the margin of the eye and generating new cells centrally to form the pre-ommatidia. It is unknown which ommatidial components are generated by the primary wave of mitoses and which are generated by the secondary pulse. Anderson (1976) suggests that in S. gregaria, cells produced by the primary wave are retinula cells, (on the grounds that growing retinula axons are seen in the optic lobe throughout the moult cycle) and the cone cells (which are identifiable at the earliest stages). Cells produced by the pulse in the maturation zone are likely to be any additional retinula cells, hair cells, or precursors of pigment cells. Cells dividing among the mature ommatidia are probably secondary pigment cells, as other components are fixed in number by this stage.

As shown in Fig. 7, there is a low level of cell division in the head epidermis adjacent to the eye margin. The pattern of mitotic activity is similar to that in the mature retina and among the maturing ommatidia (compare Fig. 7b c) and similar to patterns of mitoses in other epidermal systems (see above).

These observations lend no support to the recruitment hypothesis (Hyde, 1972). This hypothesis suggests that head-capsule epidermis undergoes a change in determination brought about by the advancing eye margin, as a result of which epidermal cells produce ommatidia. Evidence against recruitment has been presented (Nowel & Shelton, 1980) which shows that the growing eye margin never approaches bristles close to the eye margin during a succession of larval instars. If there were an increased level of cell division between the eye margin and the bristles (which may be as close to the eye margin as a single cell away) and /or a decreased level of cell death in this region (see Nowel, 1979 which shows a very low level of cell death in the epidermis during the moult cycle), a stable distance between eye margin and head epidermis could be preserved despite the occurrence of epidermal cell recruitment. The present observations on cell division in the eye and head epidermis (Nowel, 1979) lend no support to this hypothesis.

My sincere thanks go to Dr Peter M. J. Shelton for all his help, advice and encouragement while working in his lab; to Beverely Hughes for help in preparing this manuscript and for expert technical assistance; to Dr Ross for supplying our culture of lavender P. americana. We are grateful to the Science Research Council for its grant to P.M.J. Shelton.

Anderson
,
H.
(
1976
).
Postembryonic development of the insect visual system
.
Ph.D. thesis, University of Leicester
.
Anderson
,
H.
(
1978
).
Postembryonic development of the visual system of the locust, Schistocercagregaria A. Pattern of growth and developmental interactions in the retina and optic lobe
.
J. Embryol. exp. Morph
.
45
,
55
83
.
Ando
,
H.
(
1957
).
A comparative study on the development of ommatidia in Odonata
.
Sci. Rep. Tokyo Kyoiku Daig., (B)
8
,
174
216
.
Bielmann
,
G.
(
1960
).
Étude du cycles des mues chez Periplaneta americana
.
Bull. Soc. zool. Fr
.
84
,
340
351
.
Bodenstein
,
D.
(
1953
).
Postembryonic development
.
In Insect Physiology
(ed.
K. D.
Roeder
), pp.
822
865
.
New York
:
Wiley
.
Bray
,
I. S.
(
1978
).
The epidermal cell cycle of Leucophaea maderae (Blattaria)
.
Third year project, University of Leicester (unpublished
).
Eley
,
S.
(
1978
).
Postembryonic development of the insect retina - a light and electron microscope study
.
M. Phil, thesis, University of Leicester
.
Eley
,
S.
&
Shelton
,
P. M. J.
(
1976
).
Cell junctions in the developing compound eye of the desert locust Schistocerca gregaria
.
J. Embryol. exp. Morph
.
36
,
409
423
.
Flint
,
R. A.
&
Patton
,
R. C.
(
1959
).
Relation of eye color to molting in Periplaneta americana L
.
Bull. Brooklyn ent. Soc
.
54
,
140
.
Friza
,
F.
(
1928
).
Zur Frage der Farbung und Zeichnung des facettierten Insektenauges
.
Z. vergl. Physiol
.
8
,
289
336
.
Gier
,
H. T.
(
1947
).
Growth rate in the cockroach Periplaneta americana
.
Ann. ent. Soc. Am
40
,
303
317
.
Gould
,
G. E.
&
Deay
,
H. O.
(
1938
).
Biology of the American cockroach, Periplaneta americana
.
Ann. ent. Soc. Am
.,
31
,
489
498
.
Hayat
,
M. A.
(
1970
).
Principles and Techniques of Electron Microscopy: Biological Application
. vol.
1
, pp.
342
343
.
New York, London
:
Nostrand Reinhold Company
.
Hoyle
,
G.
(
1953
).
Potassium ions and insect nerve muscle
.
J. exp. Biol
.
30
,
121
135
.
Hyde
,
C. A. T.
(
1972
).
Regeneration, post-embryonic induction and cellular interaction in the eye of Periplaneta americana
.
J. Embryol. exp. Morph
.
127
,
367
379
.
Imberski
,
R. B.
(
1967
).
The effect of 5-fluorouracil on the development of the adult eye in Ephestia kühniella
.
J. exp. Zool
.
166
,
151
162
.
Karnovsky
,
M. J.
(
1965
).
A formaldehyde-glutaraldehyde fixative of high osmolality for use in electron microscopy
.
J. Cell Biol
.
27
,
137
A.
Klein
,
H. Z.
(
1933
).
Zur Biologie der amerikanischen Schabe (Periplaneta americana)
.
Z. wiss Zool
.
144
,
103
122
.
Krogh
,
A.
&
Weis-Fogh
,
T.
(
1951
).
The respiratory exchange of the desert locust (Schistocerca gregaria) before, during and after flight
.
J. exp. Biol
.
25
,
344
357
.
Locke
,
M.
(
1964
).
The structure and formation of the integument in insects
.
In The Physiology of Insecta
, vol.
3
(ed.
M.
Rockstein
), pp.
379
470
.
New York
:
Academic Press
.
Nigam
,
L. N.
(
1933
).
The life-history of a common cockroach, Periplaneta americana
.
Indian J. agrie. Sci
.
33
,
530
543
.
Nowel
,
M. S.
(
1979
).
Studies on the developing insect visual system
.
Ph.D. thesis, University of Leicester
.
Nowel
,
M. S.
&
Shelton
,
P. M. J.
(
1980
).
The eye margin and compound eye development in the cockroach: evidence against recruitment
.
J. Embryol. exp. Morph
.
60
,
329
343
.
Pantin
,
C. F. A.
(
1969
).
Notes on Microscopical Technique for Zoologists
.
London
:
Cambridge University Press
.
Ready
,
D. F.
,
Hanson
,
T. E.
&
Benzep
,
S.
(
1976
).
Development of the Drosophila retina, a neurocrystalline lattice
.
Devl Biol
.
42
,
211
221
.
Ross
,
M. H.
,
Cochran
,
D. G.
&
Smyth
,
T.
(
1964
).
Eye-color mutations in the American cockroach, Periplaneta americana
.
Ann. ent. Soc. Am
.
57
,
790
792
.
Sherk
,
T.
(
1977
).
Development of the compound eye of dragonflies (Odonata). I. Larval compound eyes
.
J. exp. Zool
.
201
,
391
416
.
Sherk
,
T.
(
1978a
).
Development of the compound eyes of dragonflies (Odonata). II. Development of the larval compound eyes
.
J. exp. Zool
.
203
,
47
60
.
Sherk
,
T.
(
1978b
).
Development of the compound eyes of dragonflies (Odonata). HI. Adult compound eyes
.
J. exp. Zool
.
203
,
61
80
.
Willis
,
E. R.
,
Riser
,
G. R.
&
Roth
,
L. M.
(
1958
).
Observations on reproduction and development in cockroaches
.
Ann. ent. Soc. Am
.
51
,
53
69
.
Yamanouti
,
T.
(
1933
).
Wachstumsmessungen an Sphodromantis bioculata Burm. V. Bestimmung der absoluten Zunahmswerte der Facettengrôsse und -anzahl (zugleigh; Aufzucht der Gottesanbeterinen. XIII. Mitteilung)
.
Anz. Akad. Wiss. Wien
.
70
,
7
8
.
1

Several studies (Klein, 1933; Nigam, 1933; Gould & Deay, 1938; Gier, 1947; Willis, Riser & Roth, 1958; Biellmann, 1960) have demonstrated the variability of the intermoult period. In the present and in other work conducted concurrently (Bray, 1978), some individuals within a group of cockroaches moult much later than the majority. There is no known way of precise staging in P. americana. The range of variability in the levels of cell division shown on particular days of the moult cycle may in part be due to differing rates of development of the animals examined at these stages.