THE MOLTING CYCLE OF THE SPINY LOBSTER, PANULIRUS ARGUS LATREILLE. II. PRE-ECDYSIAL HISTOLOGICAL AND HISTOCHEMICAL CHANGES IN THE HEPATOPANCREAS AND INTEGUMENTAL TISSUES

¹ Bermuda Biological Station and The Biological Laboratories, Harvard University and Radcliffe College, Cambridge 38, Mass.

1. The integumental tissues from the branchial region of an intermolt spiny lobster (late Stage C) are bordered by an outer and an inner integument. The outer integument consists of four layers: a thin epicuticle, the pigmented layer, the principal layer, and the membranous layer. Both the epicuticle and pigmented layer are formed before molt. The inner integument, about 1/20 the thickness of the outer integument, is composed of two layers, a very thin epicuticle and a uniformly staining endocuticle.

2. The integumental tissues are composed of columnar epidermal cells and a sub-epidermal connective tissue of a loose spongy type. Large oval reserve cells constitute by far the most predominant type of cells in this tissue. These cells store abundant amounts of carbohydrate, lipid and calcium at various stages throughout the molting cycle; they are also found in the hepatopancreas.

3. Within the tubular epithelium of the hepatopancreas are two major types of cells, secretory and absorptive. The former are swollen cells containing very large vacuoles at their distal ends. Secretion is of the apocrine type, leaving only the basal region and nucleus of the cell intact. Reconstitution of secretory cells probably occurs by regeneration from the remaining basal ends. The latter type of cells (absorptive) are tall and cylindrical with a basal or central nucleus.

4. During the intermolt period (late Stage C), a period of active feeding, there is little glycogen, phosphatase, and calcium evident in either the hepatopancreas or the integumental tissues. Lipids, however, are abundant in the epithelial tissues of the hepatopancreas.

5. External signs of an approaching molt (Stage D) become evident by the appearance of a resorptive or ecdysial line along the branchiostegites 3-4days preceding molting in the summer.

6. The external signs of an approaching molt in Panulirus reflect the more basic internal changes which occur in the integumental tissues and hepatopancreas. Growth of the outer epidermis occurs shortly after its retraction from the outer integument, 10-14 days preceding molt. Growth of the epidermis of the inside integument is accomplished through an increase in cell number at 3 days preceding molt. Resorption from the old and completion of the new inner integument occurs in a period of five days, three days preceding and two days following molt. At three days preceding molt the epidermal cells of the outer integument have already resumed an orderly alignment, are greatly elongated, and have secreted the epicuticle and pigmented layer of the new skeleton. By this time most of the principal layer of the old outer integument has been broken down by the molting fluid. The almost complete mineral and organic resorption in some areas, partial in others, allows for complete freeing of the new skeleton from the old and for further thickening of the pre-exuvial layers before ecdysis.

7. During the pre-ecdysial period (late Stage D) large numbers of reserve cells of the integumental tissues become greatly swollen, and bind or store large amounts of polysaccharide material. Glycogen is abundant in the epithelial tissues of the hepatopancreas and simultaneously accumulates in the epidermal cells of the inner integument as the latter is being formed. Available evidence (see discussion) would suggest strongly that glycogen is a necessary precursor for chitin formation. Furthermore, glycogen is perhaps intimately involved in the deposition of calcospherites in the hepatopancreas.

8. During late Stage D, phosphatase appears in the distal ends of the epidermal cells bordering the integument and in reserve cells of the connective tissue. It is suggested that phosphatase in these strategic sites of active transfer probably participates in producing molecules which are able to enter or leave the cells more readily, and in those reactions involving hydrolysis and dephosphorylation of glucose phosphate to glucose, a possible starting point for chitin formation. It is not participating in calcification of the branchial integument at this time, because calcification does not begin until the second day following molt. In the hepatopancreas, on the other hand, alkaline phosphatase is heavily concentrated at the striated border of the absorbing cells and around calcium deposition sites (calcospherites). Here the enzyme appears to be involved in important transfer and dephosphorylation reactions which occur at the surface of the absorbing cells and also to be intimately concerned with the deposition of calcium phosphate at the periphery of these same cells (see discussion).

9. Evidence available suggests that lipids, abundant in the hepatopancreas at this time, function as a major source of energy. Furthermore, the accumulation of fatty acids, glycerides, and other intermediates of oxidative as well as glycolytic metabolism during late Stage D and Stage A may contribute to the rise in osmotic pressure preceding molt, facilitating water intake. Further evidence suggests that some of the fatty acids, cholesterol and unsaponifiable fatty acids conveyed to the integumental tissues preceding molt are used in the formation of the epicuticle.

10. It is suggested that many of the organic acids, products of oxidative and glycolytic metabolism, are perhaps important carriers of calcium. Such carriers participate in conveying calcium to the hepatopancreas for storage preceding molt, and in conveying it to the integumental tissues as the skeleton progressively calcifies following molt.