Serotonin immunoreactive interneurons in the brain of the Remipedia: new insights into the phylogenetic affinities of an enigmatic crustacean taxon

Abstract

BackgroundRemipedia, a group of homonomously segmented, cave-dwelling, eyeless arthropods have been regarded as basal crustaceans in most early morphological and taxonomic studies. However, molecular sequence information together with the discovery of a highly differentiated brain led to a reconsideration of their phylogenetic position. Various conflicting hypotheses have been proposed including the claim for a basal position of Remipedia up to a close relationship with Malacostraca or Hexapoda. To provide new morphological characters that may allow phylogenetic insights, we have analyzed the architecture of the remipede brain in more detail using immunocytochemistry (serotonin, acetylated α-tubulin, synapsin) combined with confocal laser-scanning microscopy and image reconstruction techniques. This approach allows for a comprehensive neuroanatomical comparison with other crustacean and hexapod taxa.ResultsThe dominant structures of the brain are the deutocerebral olfactory neuropils, which are linked by the olfactory globular tracts to the protocerebral hemiellipsoid bodies. The olfactory globular tracts form a characteristic chiasm in the center of the brain. In Speleonectes tulumensis, each brain hemisphere contains about 120 serotonin immunoreactive neurons, which are distributed in distinct cell groups supplying fine, profusely branching neurites to 16 neuropilar domains. The olfactory neuropil comprises more than 300 spherical olfactory glomeruli arranged in sublobes. Eight serotonin immunoreactive neurons homogeneously innervate the olfactory glomeruli. In the protocerebrum, serotonin immunoreactivity revealed several structures, which, based on their position and connectivity resemble a central complex comprising a central body, a protocerebral bridge, W-, X-, Y-, Z-tracts, and lateral accessory lobes.ConclusionsThe brain of Remipedia shows several plesiomorphic features shared with other Mandibulata, such as deutocerebral olfactory neuropils with a glomerular organization, innervations by serotonin immunoreactive interneurons, and connections to protocerebral neuropils. Also, we provided tentative evidence for W-, X-, Y-, Z-tracts in the remipedian central complex like in the brain of Malacostraca, and Hexapoda. Furthermore, Remipedia display several synapomorphies with Malacostraca supporting a sister group relationship between both taxa. These homologies include a chiasm of the olfactory globular tract, which connects the olfactory neuropils with the lateral protocerebrum and the presence of hemiellipsoid bodies. Even though a growing number of molecular investigations unites Remipedia and Cephalocarida, our neuroanatomical comparison does not provide support for such a sister group relationship.

Serotonin-immunoreactive neurons in the ventral nerve cord of Remipedia (Crustacea): support for a sister group relationship of Remipedia and Hexapoda?

“…However, strong evidence from morphological studies (e.g., [14,15,22,82]) supports an ambiguous, equally likely evolutionary scenario, in which Malacostraca, Remipedia and Hexapoda are closely related. Our study presents new characters that potentially could resolve these contradicting results if extended to more taxa.…”

“…Previous neuroanatomical studies on the remipede brain [14,15,22] discovered several synapomorphies with Malacostraca and Hexapoda, such as the layout of the olfactory system including its projection neuron pathway to the protocerebrum and the architecture of the central complex. Here, we resolved the rather simple organized serotonergic system of the homonomously segmented ventral nerve cord indicating that the remipede nervous system comprises a mixture of ancestral and derived anatomical features.…”

“…Because Remipedia have many morphological characters that were considered plesiomorphic, for example the homonomously segmented trunk, paddle-like appendages and a cephalic shield, they were initially seen as the sister group to all other Crustacea or Pancrustacea in early morphological studies (e.g., [10-13]; Figure 1B). In contrast, newer comparisons of molecular sequence information together with studies of the brain anatomy, ovary structure and development changed this view and proposed Remipedia to be closely related to more derived pancrustacean lineages such as Malacostraca or Hexapoda (Figure 1C) [8,9,14-22]. In several phylogenetic analyses, Remipedia often clustered together with Cephalocarida [8,18,19,23-27].…”

“…( A ) Remipedia are composed of a cephalothorax and a homonomously segmented trunk consisting of numerous trunk somites equipped with biramous limbs (Photograph of Speleonectes tanumekes adapted from [22,66]; photograph courtesy of J. van der Ham). ( B ) Because Remipedia have many characters that are considered plesiomorphic, for example the metameric trunk, they were seen as the basal sister group to all other Crustacea in early morphological studies (adapted from [13]).…”

“…While the general morphology and the 5HT-IR of the remipede brain have already been analyzed [14,15,22], here, we describe the neuroanatomy of the ventral nerve cord focusing on the distribution and morphology of 5HT-ir neurons in three species of Remipedia. Our comparisons to already studied euarthropod taxa add new insights to the phylogenetic position of Remipedia and the crustacean sister group of Hexapoda.…”