Lateralization of the paired claws into a major crusher and a minor cutter type is determined during the 4th and 5th stages of juvenile development of the lobster, Homarus americanus. Loss of both claws during this critical period delays the determination of laterality until the 6th stage when regenerate claws are present. Continued loss into the 6th stage, and beyond, however, suppresses laterality, resulting in lobsters with paired cutter claws.

The critical period for determining claw laterality may therefore be extended for a brief time to cater for claw loss, which is especially common in these early juvenile stages.

The development of higher animals is punctuated by critical periods during which the cellular substrates of specific behaviors are established (Purves and Licht-man, 1985). The critical period, presumably, represents the optimal time for the genesis of a particular behavior and often the only time. In the lobster, Homarus americanus, such critical period concerns the development of bilateral asymmetry of its paired claws viz. claw laterality. In adult lobsters, the paired claws are elaborated into a major, crusher claw and a minor, cutter claw (Herrick, 1895). The crusher is a stout, molar-toothed claw capable of closing its thumb or dactyl only very slowly but with sufficient force to crack open the shells of bivalves such as mussels, while the cutter is a slender, incisor-toothed claw capable of closing rapidly enough to catch fish (Scrivener, 1971; Govind and Lang, 1974). These differences in behavior are due largely to the fiber composition of the claw closer muscles; the crusher muscle has exclusively slow fibers while its cutter counterpart has predominantly fast fibers (Ogonowski et al. 1980). Claw laterality is random as the crusher appears with equal probability on the right or left side (Herrick, 1895). Such randomness is due to bilateral differences in reflexive activity, which initially lateralizes the claw ganglion or CNS into a crusher and cutter side on the basis respectively of greater and lesser input (Govind and Pearce, 1986). Lateralization in the CNS is subsequently expressed in the target tissue, viz. the claws, which undergo changes in their morphology and in the fiber composition of their closer muscles. The target tissue therefore appeared essential in lateralizing the CNS; in the absence of the target tissue the CNS may not be lateralized and a crusher claw may not develop.

The critical period for the determination of laterality is during the juvenile 4th and 5th stages (Emmel, 1908; Govind and Pearce, 1989) when lobsters begin to explore the ocean floor and adopt a bottom-living or benthic habit from a previously free-floating or planktonic existence (Botero and Atema, 1982). Continual contact with the substrate on the ocean floor promotes claw activity which in turn leads to lateralization. Hence, claw laterality is determined in the CNS at a time when the claws as targets are most likely to be in use, thus demonstrating an appropriate timing in the occurrence of a critical period, which is developmentally preprogrammed, and claw activity, which is dependent on the environment.

The role of the environment was dramatically illustrated when lobsters reared in smooth-walled containers and without a substratum to manipulate failed to develop a crusher claw and instead developed paired cutter claws (Lang et al. 1978). Such a sterile environment is unlikely to occur in the wild. What is more likely to occur, however, is claw loss which is especially common in the early juvenile 4th and 5th stages (J. S. Cobb, personal communication). How has this contingency of claw loss during the critical period been met? Loss of a single claw during the critical juvenile stages accentuates the bilateral differences in activity with the result that the intact claw invariably becomes a crusher and claw literality is established (Emmel, 1908; Govind and Pearce, 1989). But what happens when both claws are lost? We report here the results of experiments when both claws were removed during the critical juvenile period. In this case, we find that the determination of laterality is delayed until the 6th stage when regenerate claws are present, thereby demonstrating an extension of the critical period for a limited time as an adaptation to claw loss.

Experiments were conducted at the Marine Biological Laboratory in Woods Hole, Massachusetts, during the summer months from June to August. Larval lobsters were obtained from the State Lobster Hatchery on Martha’s Vineyard and held communally until the molt to the 4th stage which is the beginning of juvenile development. From the 4th stage onwards, the lobsters were reared individually (Lang, 1975) in commercially molded plastic trays which provided an altogether smooth surface not easily gripped by the animals’ claws. The lobsters were reared until at least the 9th stage or later at which time the configuration of the paired claws was assessed i.e. whether they were asymmetric, consisting of a cutter and crusher, or symmetric, consisting of both cutter types (Fig. 1). For each of these conditions, a representative example was chosen in which the fiber composition of the paired closer muscles was assessed on the basis of histochemical differences in myofibrillar ATPase activity (Ogo-nowski et al. 1980). Each of the claw types has a typical staining pattern for its closer muscle (Fig. 1). The cutter muscle displays intense staining indicative of fast fibers over most of its cross-sectional area except for a lighter staining ventral band of slow fibers. The crusher muscle has a staining intensity indicative of slow fibers over its entire cross-sectional area except for a thin central band of fast fibers which transforms to slow in subsequent stages. Since these muscle patterns are consistent in past (Govind and Kent, 1982; Govind and Pearce, 1986) and present experiments, mention of a particular claw type also implies a typical closer muscle pattern in the present report.

Fig. 1.

Representative juvenile 9th stage lobsters displaying paired asymmetric claws with a left cutter and a right crusher claw (A) and paired symmetric claws of the cutter type (B). Above each claw is a typical cross-section through the mid region of the claw showing the massive closer muscle occupying 90% of the cross-sectional profile and the small, dorsally situated opener muscle. In these frozen sections, muscle fiber typing in the closer claw muscle is assessed by staining for myofibrillar ATPase activity; fast fibers with a higher specific ATPase activity stain more intensely than slow fibers. The closer muscle in the cutter claw have mostly fast fibers except for a small ventral slow band while, in the crusher claw, the muscle has almost all slow fibers except for a narrow central band of fast which in the next few molts transforms to slow. Animals ×4; cross-sections ×20.

Fig. 1.

Representative juvenile 9th stage lobsters displaying paired asymmetric claws with a left cutter and a right crusher claw (A) and paired symmetric claws of the cutter type (B). Above each claw is a typical cross-section through the mid region of the claw showing the massive closer muscle occupying 90% of the cross-sectional profile and the small, dorsally situated opener muscle. In these frozen sections, muscle fiber typing in the closer claw muscle is assessed by staining for myofibrillar ATPase activity; fast fibers with a higher specific ATPase activity stain more intensely than slow fibers. The closer muscle in the cutter claw have mostly fast fibers except for a small ventral slow band while, in the crusher claw, the muscle has almost all slow fibers except for a narrow central band of fast which in the next few molts transforms to slow. Animals ×4; cross-sections ×20.

We were able to test the hypothesis that loss of the target tissue during the critical period prevents lateralization in lobsters because of their unique ability, upon loss of a limb, to regenerate a new one. Juvenile lobsters, in particular, show a robust capacity for such regeneration. We therefore reared lobsters with a substrate and removed both claws in different stage of early juvenile development (Table 1). The paired claws were removed one day after the molt, by a gentle pinch which induced autotomy, at a preformed fracture plane without undue loss of blood. In the ensuing intermolt, limb buds form, which unfold into newly regenerated limbs at the next molt. Loss of claws in the 4th stage resulted in all lobsters developing a crusher claw and hence bilateral asymmetry, similar to their control counterparts with paired intact claws. This is explained by the fact that newly regenerated claws were present in the 5th stage which is part of the critical period for the determination of asymmetry. Removal in the 5th or 6th stage also did not suppress asymmetry because presumably paired claws were present either in the 4th or in the 4th and 5th stages. These two stages represent the critical period when the CNS is lateralized and claw laterality is established (Emmel, 1908; Govind and Rearce, 1989). It was therefore surprising to find that when claws were removed in the 4th and again in the 5th stage after they had regenerated that a significant majority of animals developed a crusher claw and hence bilateral asymmetry (Table 1). One possible explanation for these results is that the target tissue is not required for lateralization of the CNS, although this seems unlikely as previous experiments (Govind and Pearce, 1986) in which the claw was exercised showed that input from the periphery was essential in lateralizing the CNS. A more likely explanation for our results is that lateralization of the CNS was delayed to the 6th stage when regenerate claws were present. Hence the critical determinative period normally restricted to the 4th and 5th stages may be extended to the 6th stage in cases where the target tissue is missing in the earlier stages.

Table 1.

Configuration of the paired claws, whether asymmetric or symmetric, following their removal at different stages of juvenile development in lobsters reared with a substrate

Configuration of the paired claws, whether asymmetric or symmetric, following their removal at different stages of juvenile development in lobsters reared with a substrate
Configuration of the paired claws, whether asymmetric or symmetric, following their removal at different stages of juvenile development in lobsters reared with a substrate

The notion that extension of the critical period is due to claw loss may be tested bearing in mind the effects of some other experimental manipulation in determining claw laterality. For instance, lack of a substrate during the critical 4th and 5th stages prevents the development of a crusher claw resulting in paired cutter claws (Lang et al. 1978). Also loss of a single claw in the 4th or 5th stages prompts the intact claw into becoming a crusher (Govind and Kent, 1982). These experimental manipulations ought to be equally effective when applied to regenerate claws in the 6th stage, providing the determination of laterality is delayed to the 6th stage. Thus, in our next experiment, the paired claws were removed in the 4th and 5th stages and allowed to regenerate in the 6th stage. At this time, the substrate, consisting of pieces of oyster shells, was removed for the duration of the 6th stage and returned at the molt into the 7th stage. The regenerate claws lacking a substrate in the 6th stage failed to develop a crusher claw and hence bilateral asymmetry (Table 2). In another experiment, the paired claws were removed in the 4th and 5th stages and allowed to regenerate in the 6th stage. At this time, one of the paired claws, the left one, was removed and was therefore absent for the duration of this 6th stage. The results were unequivocal (Table 2), a significant majority of the lobsters developed a crusher on the right side. Clearly, both treatments for manipulating claw laterality were as effective in the regenerated claws of the 6th stage as they were in the intact claws of the 4th and 5th stages. This clearly suggests that the lateralization of the CNS, which normally occurs during the 4th and 5th stages (Emmel, 1908; Govind and Pearce, 1989) may be delayed to the 6th stage if the target tissue is missing in the earlier stages.

Table 2.

Comparison of the configuration of paired claws between control lobsters with intact claws and experimental lobsters with regenerated claws in the 6th stage subjected to a lack of substrate or unilateral claw loss

Comparison of the configuration of paired claws between control lobsters with intact claws and experimental lobsters with regenerated claws in the 6th stage subjected to a lack of substrate or unilateral claw loss
Comparison of the configuration of paired claws between control lobsters with intact claws and experimental lobsters with regenerated claws in the 6th stage subjected to a lack of substrate or unilateral claw loss

These results also show that the critical determinative period is not rigidly restricted to the 4th and 5th stages but somewhat malleabe, adjusting to claw loss by being extended to the 6th stage. Can the determination of laterality be extended indefinitely or is there a limit? These questions were answered in our final two experiments in Table 1, in which the paired claws were removed beyond the 4th and 5th stages. With the paired claws removed successively in the 4th, 5th and 6th stages, the development of a crusher claw and bilateral asymmetry became increasingly improbable as almost 50 % of the lobsters developed paired cutter claws. This number is significantly different from the control condition in which over 90% of the animals developed paired asymmetric claws. Indeed, successive claw loss from the 4th to the 7th stage almost completely suppressed the development of a crusher claw as over 85 % of the lobsters faded to develop a crusher claw and bilateral asymmetry (Table 1). Clearly, claw loss over successive juvenile stages from the 4th to the 7th can altogether prevent the lateralization of the CNS and the expression of asymmetry at the periphery.

The present experiments provide some insights into the role of the target tissue in modulating the development of the nervous system. The target tissues in this case are the paired claws whose activity provides proprioceptive and sensory input to claw ganglion. Bilateral differences in these inputs serves to lateralize the CNS such that the side with greater input becomes the crusher side while the opposite side becomes the cutter side (Govind and Pearce, 1986). Although the nature of the lateralizing process within the CNS is not known, once completed it cannot be reversed as claws that regenerate in later juveniles and adults invariably resemble their predecessors (Herrick, 1895; Kent et al. 1989). Thus lateralization is limited to a critical period encompassing the 4th and 5th juvenile stages when, coincidentally, the lobsters shift from a free-floating, planktonic (pelagic) habit to a bottom-living (benthic) habit (Botero and Atema, 1982). Such a change in life style increases the opportunity for claw activity as the claws now come into contact with the substrate on the ocean floor. The critical period for lateralizing the CNS therefore occurs at a very propitious time during primary development. Lobsters, however, can compensate for loss of the paired claws during the critical 4th and 5th stages by delaying determination to the 6th stage, thereby extending the critical period to a time when regenerate claws are present. In the absence of claws, however, lateralization of the CNS does not occur with the consequent suppression of a crusher claw and the development of paired cutter claws. The extension of the critical period underscores a form of developmental plasticity which has evolved to cater for a naturally occurring contingency viz. claw loss. It should now be profitable to study the input from the target tissue particularly as the target may be removed with impunity, thus making it possible to mimic the neural input that triggers lateralization in the CNS. In this respect, the study of claw laterality in lobsters has an advantage over studies of lateralization in song birds (Nottebohm, 1977) and mammals (Sperry, 1982) where removal of the target tissue is not as easily performed. Furthermore, because the lobster nervous system has far fewer neurons than its vertebrate counterpart, uncovering the cellular basis of laterality may be that much simpler in the lobster.

We thank R. P. Elinson and A. Wong for critical comments, M. Syslow of the Massachusetts State Lobster Hatchery for supplies of larval lobsters and the Muscular Dystrophy Association of Canada and the Natural Sciences and Engineering Research Council of Canada for financial support.

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