Histology 



J. J. GOVONI 



WHILE contemporary systematists rely upon a broad scope 

 of biological features to infer relationships among taxa, 

 the definition and comparison of morphological characters re- 

 mains one of their most useful tools. The small size and often 

 altricial development of fish larvae, however, make it difficult 

 to resolve the morphology of structures other than skeletal ele- 

 ments. By clarifying tissue composition and by enhancing mor- 

 phological resolution, histological techniques may aid the sys- 

 tematist in defining characters at the tissue as well as at the 

 microanatomical level, thereby providing additional character 

 states to be examined for synapomorphies and perhaps onto- 

 genetic precedence. Because of their small size, sections of whole 

 larvae can be prepared (Fig. 16) and structural relationships of 

 organ systems examined. Insofar as there is no clear separation 

 between gross and micro-anatomy beyond the limits of human 

 visual resolution, histological techniques may otfer yet another 

 tool useful in phylogenetic analysis. 



Techniques 



Flvi2;/o«. — Inasmuch as autolysis is rapid in larval tissue (Thei- 

 lacker, 1978), fixation is difficult (Richards and Dove, 1971). 



Specimens reared in the laboratory or specimens taken from 

 brief plankton tows (O'Connell. 1980) are the most suitable for 

 histological preparation and study; specimens sorted from field 

 collections fixed in formalin and seawater will usually yield poor 

 quality preparations. Neutral buffered (phosphate buffi;rs) for- 

 malin (see Humason, 1979) enhanced with <4% acrolein (van 

 der Veer, 1 982) is recommended for rapid and thorough fixation. 

 Glutaraldehyde (2.5%) is also a useful fixative (Hulet, 1978). 



Difference in the osmolality of tissues and ambient water may 

 distort cells and tissues, especially of marine larvae. Such arti- 

 facts have not been observed in preparations of clupeiform and 

 perciform larvae, but may be of concern in the preparation of 

 anguilliform leptocephali (Hulet, 1978). Forsterand Hong (1958) 

 and Hulet (1978) provided applicable saline solutions that may 

 eliminate distortion and enhance staining. 



Sectioning and staining. — Sxandsivd animal tissue techniques 

 (e.g., Humason, 1979)— dehydration, paraffin embedding, and 

 sectioning— have been used to trace the development of organ 

 systems (O'Connell, 1981a), as well as to assess the pathology 

 of starvation in fish larvae (Umeda and Ochiai, 1975; O'Con- 



Fig. 16. Sagiual section ot a Leiostomus xanlhurus larva, 4.4 mm notochord length (glycol methacrylate section stained with alkali blue 6B- 

 neutral red). 



Fig. 17. Example comparisons of larval fish tissue and microanatomy. Abbreviations: AM, axial musculature; CS, collagenous supporting 

 shafts; EP, epidermal cells; M, midgut; MC, mucous cell; NF, nerve fiber. (A) The integumentary epithelium of a Brevoortia patronus larva 

 showing hyaline plates (arrow), a tissue characteristic of some clupeiform larvae. Note that erosion of the outer layer of epithelium is evident. 

 (Scale bar = 20 /jm; glycol methacrylate section stained with acid fuchsin — toluidine blue.) (B) The integumentary epithelium of a Leiostomus 

 xanthurus larva showing lack of hyaline plates in epithelial cells. (Scale bar = 10 iim; glycol methacrylate section stained with alkali blue 6B — 

 neutral red.) (C) Axial musculature of a Brevoortia patronus larva showing two opposing layers of muscle fibers, a tissue characteristic of clupeiform 

 larvae. (Scale bar = 50 livn, glycol methacrylate section stained with acid fuchsin — loluidine blue.) (D) Axial musculature of a Leiostomus xanthurus 

 larva showing muscle fiber layers in parallel alignment, a tissue characteristic of perciform larvae. (Scale bar = 50 iim\ glycol methacrylate section 

 stained with alkali blue 6B — neutral red.) (E) Cross section of the elongate dorsal ray of an Echiodon dawsoni larva. (Scale bar = 20 ixm: glycol 

 methacrylate section from Govoni et al., 1984.) (F) Cross section of the elongate dorsal ray of a Bregmaceros atianticus larva. (Scale bar = 15 

 Min; glycol methacrylate section stained with acid fuchsin — toluidine blue.) 



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