Molluscan Research: Techniques for collecting, handling, preparing ...

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to physically spread the radula open. If the spreading does
not succeed, place a drop of (distilled) water on the radula
and try again. Only one part of the radula needs to be in a
good orientation without too much damage. Breaking up the
radula into segments and separating some teeth will increase
the information available (Figs 12D, E, G). For this fine
manipulation holding one’s breath helps, because ribcage
movement induces movement in the arms and hands. Mark
the radulae by drawing a circle around them with an old
needle or pair of forceps.
On glass slides, start by straightening out the radula. If
it is long, you may cut it to get a better view of tooth bases.
Small radula may be cut with two needles, crossing them to
imitate a pair of scissors. Start with the front of the radula,
because the posterior part of the radula is usually easier to
work with, as the teeth are normally not fully formed. Pull
out the front half with a needle to the edge of the water,
preferably tooth side up and at a right angle to the edge.
Quickly pull it out from the water with the needle, across the
glass and up on the glue bed. This requires some practice.
The intention is that the wet radula will soak the dry glue
enough to make it stick and firmly attach the radula. Wet
glue would invade the radula due to capillary forces.
While the radula is drying, after it has started sticking to
the substrate but before it is dry, spread out the lateral teeth,
preferably so they laterally stick to the glue. Use a needle
bent like an ice-hockey stick. You can also use a moist triple
zero paint brush, or a cat’s whisker in a holder works well for
smaller radulae. Small radulae usually stick without glue.
The position of the radula can be marked with a fine tipped
pen or dots of PVA glue.
A dry, flat-mounted radula can either be torn in the
outer part of the central field, approximately in the middle of
the ribbon, or a few rows of the radula can be cut with a
scalpel (Figs 12D, E, G) or torn with needles. Tearing a
radula can provide a better view of the basal plates of the
teeth and the teeth in the central field of Patellogastropoda.
In vetigastropods, it will often also show diagnostic teeth at
the boundary of the lateral and marginal tooth fields (lateromarginal
plate), which are typically obscured one behind the
other. Although the result of such destructive approaches is
often unpredictable, it is recommended. Curving of the
radula (e.g., by using wire mounting) will also provide good
visual access to the base of the teeth, but a physical
separation of rows by tearing the radula is usually superior.
The merits and problems of wire mounting and tearing are a
trade-off. Bradner and Kay (1995) suggested removing a few
rows at the end of the radula for a better view of the teeth but
we suggest that a tear in a more central to slightly anterior
portion is more desirable, because of wear at the very
anterior end of the radula. Manipulation of the radula other
than standard mounting is not necessary for most
neogastropods, because there is little overlap of the teeth.
Transfer the radula from the last cleaning step to a drop
of water on the chosen mounting substrate with a hooked
needle. This has to be done fast, so the radula hanging on the
needle does not dry. It dries faster in pure water than in
KOH. A very small radula may dry in two to three seconds,
GEIGER ET AL. (2007) MOLLUSCAN RESEARCH, VOL. 27
depending on aerial humidity. If it dries, the radula will stick
to the needle and may be difficult to remove, or it may
crumble more or less irreversibly.
Keep radular mounts in progress covered by an upsidedown
glass bowl to avoid dust. To handle SEM subs, use
designated stub forceps; for stubs with a peg (e.g.,
Cambridge), rather use the model for grabbing the peg, as
opposed to the rim of the stub.
Radula, histology and X-ray computer tomography
In cases where the maximum information should be
obtained from a single specimen, a conscientious choice
must be made as to which data is most important. External
morphology can be obtained from histological sections
through three dimensional reconstruction (e.g., Amira 3.1®),
but radular structures are too fine to be accurately
reconstructed; at most the major radula types (e.g.,
docoglossage, rhipidoglossate, stenoglossate) can be
identified.
The most promising method for future radular studies
seems to be X-ray computer tomography (e.g., Hagadorn et
al. 2006), but so far the resolution (ca 1 µm: http://
www.microphotonics.com/skymto.html) is too coarse for
micromolluscs. Regardless, it requires the most sophisticated
instrument of its kind plus a cyclotron and involves much
work on reconstruction.
Three-dimensional reconstruction of animals is an
exciting new avenue. Given that even basic anatomical
features are difficult to observe in dissected micromolluscs,
the only alternatives are histological serial sections with
subsequent computer assisted reconstruction. Some of us
have begun to apply these techniques to some specimens
with 1 mm shells, the animals being half that size. We show
here (Fig. 13) a section through the head region of Sinezona
rimuloides (Carpenter, 1865) and the reconstructed pairs of
odontophore cartilages, the pedal ganglia and the partially
embedded statocysts (Fig. 12). The procedures are very
labour intensive and require appropriate computer hardware,
including a graphics tablet. However, it is possible to take
cross-sections and to view the anatomy in any orientation. A
dorsal view is shown in Figure 12. We expect software
developments to make the techniques easier in their
application as well as more reasonably priced.
Optical photography
Some specimens may not be placed in the high vacuum
environment of the SEM and also, if colour is required, light
optical imaging must be employed. Two main approaches
can be pursued.
SLR camera (film or digital)
A general review on shell photography has recently
been provided elsewhere (Geiger 2006b); Häuser et al.
(2005) provided an overview on digital imaging of biological
type specimens. Marco lenses usually provide 1:1
magnification (occasionally only 0.5:1), which covers an