J. Cell Sci. 10, 79-93 (1972)
Printed in Great Britain
79
SPINDLE SHAPE CHANGES AS AN INDICATOR
OF FORCE PRODUCTION IN CRANE-FLY
SPERMATOCYTES
J. R. L A F O U N T A I N , JR.*
Department of Biological Sciences, State University of Neiv York at Albany,
Nerv York 12203, U.S.A.
Albany,
SUMMARY
Changes in spindle shape during the first and second meiotic divisions in living Nephrotoma
svturalis spermatocytes have been studied and recorded cinematographically with phase-contrast,
Nomarski differential interference, and polarization microscopy. With the Nomarski system
2 classes of discrete spindle filaments (new terminology) have been observed: continuous
filaments that extend between the poles and appear to form a cage-like framework within which
the chromosomes move, and kinetochorefilamentswhich are attached to the chromosomes at
their kinetochores and converge toward the poles. Spindle deformation occurs in a manner
consistent with the hypothesis that whatever tensile forces are transmitted by the kinetochore
filaments are balanced by compressive forces acting on the cage-like framework of continuous
filaments causing them to become splayed. Just when these forces would be expected to reach
a maximum, kinetochore filaments undergo a noticeable increase in contrast. It has not been
determined whether this change represents a crowding of linear elements or addition of dry
mass to a constant number of linear elements.
INTRODUCTION
The occurrence of dimensional changes (i.e. prometaphase elongation, metaphase
shortening, and anaphase elongation) of mitotic spindles at specific times during the
division process has been described for several different types of cells (Duncan &
Peridsky, 1958; Jacquez & Biesele, 1954; Taylor, i960), but little functional significance has been ascribed to these changes of shape. Very striking deformations occur
in crane-fly spermatocyte spindles, which are clearly demarked by a mitochondrial
sheath, and anaphase movement is preceded by a noticeable spindle shortening and
widening (Dietz, 1969). The present study was undertaken to document these dimensional changes in spermatocyte spindles of the crane fly, Nephrotoma suturalis, and
determine their probable cause.
The Zeiss/Nomarski differential interference equipment for transmitted-light
microscopy was used extensively in this study to render visible small gradients in
optical path, such as those produced by linear elements of the mitotic spindle (Allen,
David & Nomarski, 1969). This study represents the first detailed investigation of
an animal spindle with the Nomarski system under conditions comparable to those
used in studies on Haemanthus endosperm by Bajer & Allen (1966a, b).
• Present address: Laboratorium fur Elektronenmikroskopie, Institut fiir Allgemeine
Botanik, Eidgendssische Technische Hochschule, Zurich, Switzerland.
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J. R. LaFountain, Jr.
MATERIALS AND METHODS
Larvae of Nephrotoma suturalis were cultured in the laboratory on macerated tissue paper
and ground nettle leaves according to the method of Forer (1964). Spermatocytes from testes
of fourth instar larvae were spread as a monolayer under KEL-F 10 oil on to a large (24 x
50 mm) coverglass, which was inverted on to a smaller (22 x 40 mm) coverglass to make a
sandwich and sealed with Valap (a mixture of Vaseline, lanoline and paraffin). A ring of Vaseline
was applied to the smaller coverglass before making the sandwich, and it served as a spacer to
reduce cell flattening. The coverglass sandwich was then mounted on to a brass holder that fits
on to a microscope stage. Spermatocytes prepared in this way could live for 4-5 h at room
temperature (20—23 °C), and both meiotic divisions were observed in many cells.
Microscopical observations were made with a Zeiss Photomicroscope I equipped with selected
high-extinction Nomarski differential interference and polarization optics and positive phasecontrast optics. A neofluar (40/0-75 N.A.) objective was used for phase contrast; strain-free
(40/085 N.A.) and achromatic (40/085 N.A. oil immersion) objectives were used with polarization optics; and an achromat (40/0-85 N.A. oil immersion) and a planachromat (100/125 N.A.
oil immersion) were used with the Nomarski system. Tungsten and mercury arc (with a green
(546 run) filter) lamps were used with phase and Nomarski optics, and a Spectra-Physics argonion laser (LaFountain, Muckenthaler, & Allen, 1968) was used as a light source for the polarization microscope. Operational extinction factors for the Nomarski equipment ranged between
200 and 250, while extinction factors for the polarization optics were between 5 x io 3 and
8 x io 3 .
Cin6 records were made on Eastman Recordak Micro-file AHU, Plus X Negative, and Ektachrome Commercial films with an Arriflex 16-mm camera driven by a Sage series 500 Cin6photomicrographic apparatus; 3 5-mm records were made with the Photomicroscope I automatic
camera on Adox KB 14 film. Black and white films were processed in Diafine and colour films
were processed commercially.
RESULTS
Nephrotoma spermatocytes are cultured so that the spindles are not flattened but
have a circular cross-section in a plane perpendicular to the pole-to-pole axis. Spindle
length is defined as the distance between the 2 poles of the spindle, and the accuracy
of measurements of spindle length depends on the accuracy with which the poles can
be determined. Since the spindle poles are defined in these cells as isotropic regions
at the polar ends of the highly birefrigent chromosomal fibres, comparisons of interpolar distances at different stages of division can be made with the polarization microscope (Fig. 1). Spindle width (at the spindle equator) can be determined accurately
because the spindle border is defined at its periphery by a dense mitochondrial
sheath, which completely encases the spindle (Figs. 2, 3).
In order to translate spindle shape changes as viewed in the plane of the pole-topole axis into shape changes of the entire spindle, it is necessary to know if the shape
of the spindle is constant about the pole-to-pole axis. If spindles are axially symmetrical, then the shape of the entire spindle is defined by its longitudinal median section.
As a test of spindle symmetry, ocular micrometer measurements of spindle equatorial
width (perpendicular to the optical axis) were compared with measurements of spindle
width parallel to the optical axis made with the calibrated fine focus of the Photomicroscope I. For these measurements Nomarski optics were used to 'optically section' (Allen et al. 1969) cells and thereby determine the vertical distance from the
upper boundary of the spindle to the lower boundary as defined by the mitochondrial
Spindle shape changes and force production
81
sheath. Over 20 measurements were made of cells during different stages of division
in several preparations and were restricted to spindles in which both poles were clearly
in the same focal plane. In all cases studied, spindle equatorial width along both axes
was the same (the 2 values differed from one another by no more than 1 /tm), indicating
that the spindles were not flattened by the preparation method used and that an
assumption of axial symmetry along the pole-to-pole axis is valid.
During meiosis I in Nephrotoma spermatocytes, a spindle begins to form between
the 2 asters immediately after the nuclear membrane breaks down, and the chromosomes generally begin to undergo random metakinetic movements and chromosome
stretching. (A full description of these and other chromosome movements will not be
included here because they closely resemble phenomena observed in other species of
crane flies, which have been described in great detail by Dietz, 1956.) As the spindle
forms, the asters move in opposite directions and establish the spindle poles (Fig. 2 A,
B). Some structural rigidity on the part of the spindle can be inferred from the fact
that cells elongate parallel to the spindle axis, and also, neighbouring cells may be
pushed aside to allow for this elongation. Within 15-20 min after nuclear membrane
breakdown, the spindle appears ellipsoidal and has the shape of a prolate spheroid
(Fig. 2B).
As the chromosomes become stabilized into a metaphase plate configuration, the
spindle itself shortens, while its equatorial diameter increases. Micrographs made
with either phase-contrast or Nomarski optics illustrate this change (Fig. 2 c). Moreover, time-lapse cine records made with the polarization microscope permit an
accurate determination of the extent of spindle shortening (Fig. 1). In the most
extreme cases, interpolar distance is reduced as much as 20% during metaphase as
compared to early prometaphase. Just before the onset of anaphase, spindles appear
nearly diamond-shaped in optical section, and their shape approximates that of a
bicone (Fig. 2 c). After separation of sister dyads and during anaphase, spindle shape
gradually changes once more. The diamond shape of metaphase slowly changes into
an ellipsoid, and the shape of the spindle returns to a prolate spheroid (Fig. 2E).
Spindle width decreases, and spindle length slowly increases (Fig. 1), so that in many
instances, spindle shape at the moment when dyads reach the poles is almost identical
to the prolate spheroid observed during prometaphase. The main observable difference
between the 2 stages is the position of the chromosomes within the spindle. Although
not all spindles studied showed anaphase elongation back to prometaphase length,
none showed interpolar distances greater than prometaphase values until after cytokinesis begins (Fig. 2F).
Spindles in secondary spermatocytes of Nephrotoma show shape changes similar to
those observed in meiosis I (Fig. 3). Just before the monads are transported to the
poles, the equatorial diameter of secondary spindles reaches a maximum and the
length is reduced. During anaphase, spindle shape gradually changes to a prolate
spheroid. Also, as was the case in meiosis I, spindle length does not increase to more
than what it was during prometaphase until after cytokinesis ensues.
Exploration of Nephrotoma spermatocyte spindles with the Nomarski system has
revealed several new details about the linear elements in these spindles. Both primary
82
J. R. LaFountain, Jr.
and secondary spindles contain numerous longitudinal fibrils which are visible as
refractile bodies (Figs. 2-7) with a diameter of 0-2-0-3 / t m - (If t n e reader is unfamiliar
with the interpretation of differential interference images, an earlier description of the
structure and organization of the mitotic spindle in Haemanthus endosperm (Bajer &
Allen, 1966 a), as well as a description of the Nomarski system (Allen et al. 1969),
should be consulted for details.) These elements can be observed extending from the
kinetochore regions of the chromosomes toward the poles and also extending between
the poles without any apparent connexions to the chromosomes. Because these elements are not similar in either size or distribution to the longitudinal spindle elements previously observed with the light microscope (i.e. birefringent spindle fibres
described by Forer, 1966, 1969), the term spindle filament has been applied to
these newly revealed structures to mark this distinction. Spindle filaments that
extend from the kinetochore regions of the bivalents and univalents to the poles are
called kinetochorefilaments,and filaments that extend between the poles and do not
terminate at kinetochores are called continuous filaments.
During the first maturation division, numerous spindle filaments can be seen in
Nephrotoma spindles soon after the nuclear membrane breaks down, and within a few
minutes after the onset of spindle formation, the entire spindle region becomes
packed with filaments. Optical sections through the spindle during prometaphase
demonstrate that continuous filaments appear somewhat arced (Fig. 2B) and form
a cage-like framework within which the chromosomes move. It is possible in some
cells to trace single continuous filaments from one pole to the other. However, due to
the slight curvature of these elements, micrographs of single filaments extending from
pole to pole are not available. Kinetochore filaments are connected to the bivalent
chromosomes at the kinetochores and extend toward the poles. Although kinetochore
filaments have not been traced all the way to the poles, they can be traced very close
to the poles (Fig. 5 A), and they are distributed one per autosomal chromatid (Figs. 5 A,
6 A). Thus, birefringent chromosomal fibres of sister dyads are made up of 2 kinetochore filaments (Fig. 5). In the cases of unpaired sex univalents, each chromatid has
2 kinetochore regions with a kinetochore filament radiating from each (Fig. 6B).
Although kinetochore filaments are distributed 2 per univalent chromatid, a ratio
of 2 kinetochore filaments per chromosomal fibre is maintained.
As metaphase approaches and the shape of the spindle gradually changes to a
bicone, the distance between the centrally located chromosomes and the mitochondrial
sheath generally increases, and some of the more peripheral continuous filaments
splay out in this region between the chromosomes and the mitochondrial sheath
(Fig. 7). In some instances the appearance of the continuous filaments suggests that
they actually bend at the spindle equator (Fig. 4). During this time the visibility of,
or contrast generated by, kinetochore filaments becomes markedly enhanced over
that during prometaphase (Fig. 7). This increased contrast takes place without any
apparent change in filament thickness. During anaphase, kinetochore filaments become shorter and remain distinct from continuous filaments (Fig. 4). The kinetochore
filaments of the lagging sex univalents are easily distinguished from continuous and
perhaps interzonal filaments because they maintain a state of enhanced contrast until
Spindle shape changes and force production
83
and during the time they are transported to the poles during telophase (Fig. 2D, E).
(Interzonal fibres are clearly visible at the level of the light microscope (Schrader,
i953;Bajer, 1968; Forer, 1969). At present, however, one cannot exclude the possibility
that these are segments of continuous filaments, the courses of which could have been
changed within chromosomes.)
The arrangement of spindle filaments in secondary spermatocytes closely resembles
that found in meiosis I (Fig. 3). Continuous filaments appear to be distributed throughout the spindle forming a kind of longitudinal framework, while kinetochore filaments
are attached to the chromosomes at the kinetochores. Enhanced contrast in kinetochore
filaments can also be observed before and during anaphase, but the appearance of this
phenomenon is not as striking as in meiosis I.
DISCUSSION
Regarding the mechanism of chromosome transport during cell division, it is generally believed that poleward-pulling forces applied to the kinetochores are responsible
for anaphase movement (for review: Forer, 1969; Nicklas, 1970). Chromosomal fibres
undoubtedly are involved in either generating or transmitting the tensile forces that
result in movement (Forer, 1966, 1969). Since the spindle poles approach one another
to a limited extent, the spindle must contain supporting elements that resist these tensile forces. The presence and arrangement of continuous or interpolar microtubules in
many plant and animal spindles suggests that the role of these elements is to provide
the supporting framework within which chromosomes move (Bajer, 1968; Nicldas,
1970; Nicklas & Staehley, 1967). The findings obtained here in living crane-fly meiotic
spindles offer additional evidence that continuous elements provide the structural
framework needed to support the intraspindle forces that bring about chromosomal
movement. In addition, the appearance and distribution of continuous filaments in
Nephrotoma spindles, coupled with the dramatic shape changes of the entire spindle
during the course of division, are compatible with a simple hypothesis regarding the
development and deployment of forces within the spindle from prometaphase through
anaphase. That is, tensile forces between the chromosomes and poles develop long
before anaphase movement commences, and the different stages of division (prometaphase, metaphase, and anaphase) are results of continued action of the same mechanism.
Accordingly, metaphase is a state of equilibrium when the forces counterbalance
one another (Ostergren, 1945, Ostergren, Mole-Bajer & Bajer, i960; Mclntosh,
Hepler & VanWie, 1969).
The occurrence of active chromosome movement and chromosome stretching
during prometaphase in Nephrotoma spermatocytes suggests that tensile forces acting
between the chromosomes and poles are already active at that time (Dietz, 1956;
Nicklas, 1970). In early prometaphase continuous filaments are distributed more or
less uniformly throughout the spindle and some of the more peripheral ones appear
slightly arced. As prometaphase proceeds and the chromosomes became stabilized
on to the spindle equator, the outer continous filaments, lying close to the periphery
of the spindle, tend to splay out, or bend to a greater degree, causing the spindle to
6-2
84
J- R- LaFountain, Jr.
widen and assume the shape of a bicone. These changes can be explained in terms of
increased compression applied to continuous filaments due to increased tensile forces
between the chromosomes and the poles. As tension toward one pole is balanced by
forces pulling in the opposite direction, the chromosomes equilibrate at a point half
way between the 2 poles. The result of this is metaphase, during which time spindle
deformation reaches a maximum, and there is no net chromosome movement to the
poles.
According to this interpretation, it would be expected that tension on the kinetochore filaments and the balancing compression on continuous filaments would both
diminish upon separation of sister dyads and the onset of anaphase. The results bear
this out. When the chromosomes move to the poles during anaphase, spindle width
decreases and the continuous filaments become less splayed (Fig. 4). Also, interpolar
distance generally increases (Fig. 1), but never beyond the maximum established
during prometaphase. Spindle elongation during anaphase may then be interpreted
as a result of the release of continuous filaments from compression. If this entire
interpretation is correct, then the shape of the spindle may be a sensitive indicator of
the magnitude of the balanced forces acting within the spindle.
Included in this interpretation is an assumption that the apparent contraction of
chromosomes during metaphase is not a result of a reduction in the tensile forces
applied to them (as was suggested by Dietz, 1956) but is due to force-independent
changes within the chromosomes themselves. Since chromosome condensation occurs
during other stages (i.e. prophase and telophase) without the influence of motive
forces, it is concluded that the above assumption calling for self-condensation is valid.
The discovery of both continuous and kinetochore filaments as refractile bodies in
Nephrotoma spindles has raised several interesting questions. First, what is the
relationship between spindle filaments and microtubules? That is, are spindle filaments composed of several microtubules (Bajer & Jensen, 1969), and if so, how many
microtubules are there per filament? Secondly, what is the relationship between
spindle filaments and birefringent spindle fibres seen with the polarization microscope?
Continuous filaments can be found in regions where little or no birefringence can be
detected (the interzone (see Forer, 1969) and the more peripheral regions of the
spindle between the chromosomes and the mitochondrial sheath). These findings plus
other data suggest that the interpretation of birefringence solely in terms of oriented
filamentous material may not be acceptable (J. R. LaFountain, in preparation).
Thirdly, what is the significance of the marked increase in contrast generated by kinetochore filaments during the later stages of prometaphase through anaphase? Since
the steepness of the shadows cast across linear elements with the Nomarski system is a
function both of the gradient in optical path difference in the object and the instrumental bias retardation (Allen et al. 1969), changes in the contrast of an object at constant bias retardation reflect size and/or refractive index changes. If kinetochore
filaments change in size at this time, the change must be a subtle one. In order to
account for an increase in contrast, the elements comprising the spindle filament
would have to become more condensed, thus increasing the mean refractive index in
the filament area. Alternatively, the same effect could be achieved by the addition of
Spindle shape changes and force production
85
dry mass to the filament. One might speculate whether some undetected' last elements'
(e.g. microtubule cross-bridges, additional microtubules, microfilaments, etc.) of the
force-producing mechanism might be laid down at this stage and contribute sufficient
dry mass to be observed as enhanced contrast.
Despite the extensive study of Behnke & Forer (1966) on Nephrotoma spindles, our
knowledge of spindle ultrastructure in these cells is very incomplete. In order to
answer any of the questions raised above, a more thorough investigation of the
ultrastructure of Nephrotoma spindles must be made. Studies toward this end are now
being conducted.
The author thanks Dr Robert D. Allen for his advice and criticism during the course of this
investigation; Dr F. A. Muckenthaler for help during the early stages; and Robert Speck and
Dale Rice for technical assistance.
This work was supported by a predoctoral fellowship from the National Institute of Health
(1-F1-GM-40, 135-01) and Program Project Grant G M 08691 from the National Institute of
General Medical Sciences administered by Dr R. D. Allen.
REFERENCES
ALLEN, R. D., DAVID, G. B. & NOMARSKI, G. (1969). The Zeiss/Nomarski differential inter-
ference equipment for transmitted-light microscopy. Z. iviss. Mikrosk. 69, 193-221.
BAJER, A. (1968). Behavior and fine structure of spindle fibers during mitosis in endosperm.
Chronwsoma 25, 249—281.
BAJER, A. & ALLEN, R. D. (1966a), Structure and organization of the living mitotic spindle of
Haemanthus endosperm. Science, N.Y. 151, 572-574.
BAJER, A. & ALLEN, R. D. (19664). Role of phragmoplast filaments in cell-plate formation.
J. Cell Sci. 1, 4SS-462.
BAJER, A. & JENSEN, C. (1969). Detectability of mitotic spindle microtubules with the light and
electron microscopes. J. Microscopie 8, 343-354.
BEHNKE, O. & FORER, A. (1966). Some aspects of microtubules in spermatocyte meiosis in a
crane fly (Nephrotoma suturajis, Loew): Intranuclear and intrachromosomal microtubules.
C. r. Trav. Lab. Carhberg 35, 437-455DIETZ, R. (1956). Die Spermatocytenteilungen der Tipuliden II. Graphische Analyse der
Chromosomenbewegung wahrend der Prometaphase I im Leben. Cliromosoma 8, 183-211.
DIETZ, R. (1969). Bau und Funktion des Spindelapparatus. Naturwissenschaften 56, 237-248.
DUNCAN, R. E. & PERIDSKY, M. D. (1958). The achromatic figure during mitosis in maize
endosperm. Am. J. Bot. 45, 719-729.
FORER, A. (1964). Evidence for Two Spindle Fiber Components: A Study of Chromosome Movement in Living Crane-fly (Nephrotoma suturalis) Spermatocytes using Polarization Microscopy
and an Ultraviolet Microbeam. Doctoral Thesis, Dartmouth College.
FORER, A. (1966). Characterization of the mitotic traction system and evidence that birefringent
spindle fibers neither produce nor transmit force for chromosome movement. Cliromosoma
19, 44-98.
FORER, A. (1969). Chromosome movements during cell-division. In Handbook of Molecular
Cytology (ed. A. Lima-de-Faria), pp. 553-601. Amsterdam: North-Holland Publishing Co.
JACQUEZ, J. A. & BIESELE, J. J. (1954). A study of Michel's film on meiosis in Psophus stridulus,
L. Expl Cell Res. 6, 17-29.
LAFOUNTAIN, J., MUCKENTHALER, F. & ALLEN, R. D. (1968). Argon-ion laser as a source for
physical microscopy. Biophys. J. 8, A159.
MCINTOSH, J. A., HEPLER, P. K. & VANWIE, D. G. (1969). A model for mitosis. Nature, Lond.
224, 659-663.
NICKLAS, R. B. (1970). Mitosis. In Advances in Cell Biology, vol. 2 (ed. D. M. Prescott, L.
Goldstein & E. H. McConkey). New York: Appleton-Century-Crofts.
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R. B. & STAEHLEY, C. A. (1967). Chromosome micromanipulation. I. The mechanism of chromosome attachment to the spindle. Chromosoma 21, 1-16.
OSTERGREN, G. (1945). Equilibrium of trivalents and the mechanism of chromosome movement.
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NICKLAS,
OSTERGREN, G., MOLE-BAJER, L. & BAJER, A. (i960). An interpretation of transport phenomena
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SCHRADER, F. (1953). Mitosis, Hit Movement of Chromosomes in Cell Division, 2nd edn. New
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E. W. (i960). Dynamics of spindle formation. Ann. N.Y. Acad. Sci. 90, 430-434.
TAYLOR,
(Received 8 January 1971—Revised 21 June 1971)
Fig. 1. Three different Nephrotoma suturalis spermatocytes during meiosis I with
polarization microscopy (40/085 N.A. oil-immereion achromatic objective and laser
illumination): A-F, first cell from early prometaphase through anaphase showing
prometaphase elongation (A, B), metaphase shortening (c, D), and anaphase elongation
(E, F); G-I, second cell from prometaphase (G, H) to metaphase (1); J-L: third cell
during metaphase (j) and anaphase (K, L). Birefringent bands in the interzone are
the chromosomal fibres of the lagging univalent sex chromosomes (u). Arrows locate
poles. Spindle fibre birefringence appears as either positive (white) or negative
(black) contrast depending on compensator setting.
Spindle shape changes and force production
J. R. LaFountain, Jr.
Fig. 2. First meiodc division in a Nephrotoma primary spermatocyte with Nomarski
differential interference optics (100/125 N.A. oil-immersion planachromatic objective) and mercury arc illumination: A, late diakinesis; B, prometaphase; c, metaphase;
D, anaphase; E, telophase; F, cytokinesis, cf, continuous filament; cif, continuous or
interzonal filament; d, dyad; kf, kinetochore filament; m, mitochondrial sheath;
t, tetrad; u, univalent sex chromosome.
Spindle shape changes and force production
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J. R. LaFountain, Jr.
Fig. 3. Second maturation division in a Nephrotoma secondary spermatocyte with
Nomarski differential interference optics (100/125 N.A. oil-immersion planachromatic objective): A, interphase; B, prometaphase; c, late prometaphase; D, metaphase; E, anaphase; F, anaphase; c, anaphase, H, telophase; 1, cytokinesis, cif, continuous or interzonal filament; d, dyad; kf, kinetochore filament; m, mitochondrial
sheath; mo, monad.
Spindle shape changes and force production
4A
Fig. 4. First maturation division with Nomareki differential interference optics
(100/1-25 N.A. oil-immersion planachromatic objective); A, prometaphase, both
continuous and kinetochore filaments visible; B, early anaphase, kinetochore filaments accentuated and peripheral continuous filaments {cf) splayed; c, later anaphase,
continuous and perhaps interzonal filaments visible and peripheral continuous filaments (cf) less splayed, cf, continuous filament; cif, continuous or interzonal filament;
kf, kinetochore filament.
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J. R. La Fountain, Jr.
Fig. 5. A Nephrotoma primary spermatocyte with both A, Nomarski optics (100/
1 25 N.A. oil-immersion planachromatic objective) and B, polarization optics (40/
085 N.A. oil-immersion achromatic objective and mercury arc illumination). The
birefringent chromosomal fibre (arrow) in B contains 2 kinetochore filaments (arrow
in A).
Fig. 6. A, Nephrotoma primary spermatocyte during metaphase with Nomarski optics
(100/125 N.A. oil-immersion planachromatic objective); kinetochore filaments are
distributed one per autosomal chromatid, or 2 per dyad (arrow). B, Univalent sex
chromosomes remain at the spindle equator during autosomal anaphase I and have
2 kinetochores per chromatid with a kinetochore filament radiating from each kinetochore (arrow). Nomarski optics (100/1-25 N.A. oil immersion planachromatic
objective).
Fig. 7. A Nephrotoma primary spermatocyte with Nomarski optics (100/1-25 N.A.
oil-immersion planachromatic objective): A, prometaphase; B, metaphase. Kinetochore filaments show enhanced contrast during metaphase (compare contrast in
filaments in A (arrow) with contrast in B) and peripheral continuous filaments become
splayed (paired arrows in B).
Spindle shape changes and force production