The male gamete of
Equisetum
is the largest and structurally most complex of those so far known in living pteridophytes. The ultrastructure of the mature gametes, is described with particular reference to the influence of the multilayered structure (MLS) on its form. In
Equisetum
this organelle comprises a band of over 300 microtubules, underlain along its anterior edge by a lamellar strip, 15-20 µm in length, and forming a sinistral spiral of 2 1/2 gyres. The tubules extend from the strip, at an angle of about 40°, to form a broad sheath around the twisted pyriform nucleus located in the posterior half of the cell. From the anterior tip of the lamellar strip to the posterior end of the nucleus the gamete completes a helix of 3 1/2 gyres, traversed throughout by the microtubular band. As a result of growth of this band during spermatid metamorphosis, and the 40° angle between the elates of the lamellar strip and the microtubules, the strip is displaced anteriorly and laterally relative to the nucleus. In the mature gamete, although the strip and the nucleus remain interconnected by the microtubular band, only the posterior half of the strip lies directly above the anterior third of the nucleus. The precise interrelationship between nucleus and MLS is illustrated by reconstructions which display the spermatozoids as they would appear if uncoiled. The 80-120 flagella are inserted outside that part of the micro tubular band lying anterior to the nucleus. Their basal bodies retain the proximal cartwheel and stellate transition regions found already in spermatids, but in the mature gametes they are invested with collars of osmiophilic material. The axonemes depart at 10° tangentially from the helix and extend backwards parallel with the tubules of the microtubular band. In consequence of the overlapping gyres of the helix the flagella lie in a spiral groove, similar to that found in cycad spermatozoids. From this groove the plasma membrane closely follows the external surface of the microtubular band. Contrasting with other archegoniates, maximal structural differentiation of the MLS is found in the mature spermatozoid. Flat-bottomed keels are present on the microtubules overlying the lamellar strip in which three distinct strata can be recognized. The two outer, consisting of alternating plates of electron-opaque and electron- transparent material, are separated by a continuous electron-opaque sheet. The innermost stratum comprises a continuous layer of finely granular material. Overlying the external anterior rim of the microtubular band is an osmiophilic crest. This retains the regularly banded substructure found in spermatids, but in mature spermatozoids is far more prominent than at any other time during spermatogenesis. It contains an electron-transparent lumen and is continuous with both the anterior ends of the microtubules and the anteriormost lamellar plates. Between the inner gyres of the MLS the crest is confluent with extensive sheets of smooth endoplasmic reticulum. Underlying the lamellar strip is a spiral mitochondrion with prominent dilated cristae. The central cytoplasm contains at least 100 pleomorphic mitochondria, together with from 15 to 25 amyloplasts and a few microbodies. In the nucleus, in addition to condensed chromatin, are several spherical electron-opaque bodies and aggregations of membrane-bound vesicles. Structures identical in appearance with the former also occur in the cytoplasm, and it is suggested that they may be nuclear in origin, as are similar bodies in animal spermatogenesis. The vesicles may represent portions of redundant nuclear envelope whose extrusion into the cytoplasm was prevented by the ensheathing microtubular band. Pores are still present in the nuclear envelope, where this is not invested by the band. The mature spermatozoids are liberated from antheridia within mucilaginous sacs bounded by fibrillar cell wall material, thought to contain lipid droplets promoting their dispersal when in contact with water. On escaping from the sacs the spermatozoids elongate slightly, and profiles of disrupted flagella are frequently encountered. Occasionally the microtubular band ensheathing the posterior part of the nucleus also becomes disorganized. There is no evidence of the utilization of amyloplast starch as an energy source during motility, and, in contrast to ferns and bryophytes, there is no sequestration of the central cytoplasm by the swimming spermatozoids.
Individual microtubules viewed by immunofluorescence and electron microscopy in the same PtK2 cell