Effects of tentacle amputation and regeneration on the morphology and activity of the olfactory center of the terrestrial slug Limax valentianus

Abstract

SUMMARYThe tentacles of pulmonates regenerate spontaneously following amputation. The regenerated tentacle is equipped with all the elements necessary for normal olfactory functioning, and the slugs can behave as well as they did before the tentacle amputation. However, it is not known what changes occur to the olfactory center procerebrum in the brain at the morphological and physiological levels. Here, we investigated the innervation of tentacular nerves into the procerebrum by examining the size of the terminal mass (input layer from tentacular nerves) of the procerebrum and also by staining afferent nerves immunohistochemically at 15, 58 and 75 days following unilateral amputation of the superior and inferior tentacles. The size of the terminal mass was significantly decreased, and the Phe-Met-Arg-Phe-NH 2 ergic (FMRFamidergic) afferent nerves disappeared by 15 days following the tentacle amputation. However, the size of the terminal mass had recovered substantially by 58 days, as the tentacle regenerated. The FMRFamidergic innervation into the cerebral ganglion was also restored by this time. An extended recovery (75 days), however, did not result in any further increase in the size of the terminal mass. We also recorded the local field potential (LFP) oscillation in the procerebrum. We found that the oscillatory frequency of the LFP had decreased at 15 days following the tentacle amputation but had recovered at 58 and 75 days. These results suggest that the amputation and regrowth of the tentacle are accompanied by the respective degeneration and re-innervation of olfactory nerves, and these changes in the innervation status affect the basal state of LFP oscillation.

Olfactory Memory Storage and/or Retrieval Requires the Presence of the Exact Tentacle Used During Memory Acquisition in the Terrestrial SlugLimax

“…5). Such morphological recovery seems to underlie the ability of slugs with only a pair of regenerated ST (Matsuo et al, 2010b) or pairs of regenerated ST and IT (Fig. 3), to accomplish an odor-aversion learning task.…”

“…In a previous study, we demonstrated that the amputated tentacles recover their normal olfactory functions within 34 days (Matsuo et al, 2010b). We thus tested whether the olfactory memory can be retained for a long enough time to allow the amputated tentacles to regenerate.…”

“…Previously, we have demonstrated that the slugs can acquire and retrieve olfactory memory using only regenerated superior tentacles (Matsuo et al, 2010b). We here explored whether the slugs can acquire and retrieve the memory using two pairs of regenerated tentacles that had been amputated 34 days prior to the conditioning.…”

“…Because the amputated tentacles are able to regenerate and regain their function following a recovery period (Chase and Kamil, 1983;Matsuo et al, 2010b;, in the present study we sought to exploit this regenerative ability to examine whether or not the same tentacles must be used during the acquisition and retrieval phases in order for the memory to be retrievable.…”

Regeneration in spiralians: evolutionary patterns and developmental processes

“…However, across the phylum, a number of structures have been shown to regenerate. These include the foot, anterior neural elements, tentacles, and even the entire head of some gastropods (Moffett, 1995(Moffett, , 1996Gorbushin et al, 2001;Matsuo et al, 2010;Tuchina and Meyer-Rochow, 2010;Hoso, 2012), the siphon and parts of the shell and mantle of some bivalves (Ansell et al, 1999;Tomiyama and Ito, 2006;Nuñez et al, 2013), and the arms, tentacles, and suckers of many cephalopods (Feral, 1978;Bush, 2012;Fossati et al, 2013) including those of giant squid (Aldrich and Aldrich, 1968). Greater sampling of regenerative ability is needed within each of these groups, as well as in the mollusc lineages in which there are no data on regeneration, in order to obtain a clearer picture of the distribution of regenerative ability across this phylum.…”