DOI: 10.5061/dryad.tqjq2bw3x
title: Supplementary information for: Multisensory integration by polymodal sensory neurons dictates settlement in actinulae larvae

description: Multisensory integration (MSI) combines information from more than one
 sensory modality to elicit behaviors distinct from unisensory behaviors.
 MSI is best understood in animals with complex brains and specialized
 centers for parsing sensory information, but the dispersive larvae of
 sessile marine invertebrates utilize multimodal environmental sensory
 stimuli to base irreversible settlement decisions on, and most lack
 complex brains. Here, we examined the sensory determinants of settlement
 in actinula larvae of the hydrozoan Ectopleura crocea (Cnidaria),
 which possess a diffuse nerve net. A factorial settlement study revealed
 that photo-, chemo-, and mechano-sensory cues each influence the
 settlement response, which was complex and dependent on specific
 combinations of cues, therefore indicating MSI. Mechanosensory cues either
 inhibited or enhanced settlement rates depending on the presence or
 absence of chemical and light cues in the environment. Sensory gene
 expression over development peaked with developmental competence to
 settle, which in actinulae, requires cnidocyte discharge. Transcriptome
 analyses also highlighted several deep homological links between cnidarian
 and bilaterian mechano- chemo- and photo-sensory pathways. Fluorescent in
 situ hybridization studies of candidate transcripts suggested cellular
 partitioning of sensory function among the few cell types that comprise
 the actinula nervous system, where ubiquitous polymodal sensory neurons
 with putative chemo- and photo-sensitivity interface with mechanoreceptive
 cnidocytes. We propose that a simple multisensory processing circuit,
 involving polymodal chemo/photosensory neurons and mechanoreceptive
 cnidocytes, is sufficient to explain MSI in actinulae settlement. Our
 study demonstrates that MSI is not exclusive to complex brains, but likely
 predated and contextualized their evolution.
description: Please see the README document
 (README.md). This dataset includes additional supplementary data for the
 article: Birch S., and Plachetzki D. 2023. Multisensory integration by
 polymodal sensory neurons dictates settlement in actinulae larvae.
 Specifically, we include the raw behavioral data from the larval
 settlement study, the pilot study data used to determine the
 mechanosensory condition in the larval settlement study, and the RNA
 fluorescent in-situ hybridization (FISH) probe sequences used in Birch and
 Plachetzki (2023). Below,
 you will find the methods for the mechanosensory pilot study and
 information on the target sequences for RNA FISH probes. For information
 on the main larval settlement study see Birch and Plachetzki
 (2023). Actinula larvae mechanosensory pilot
 study Colonies of <em>E. crocea</em> were
 collected by hand at two sites, the UNH CML pier in New Castle, NH, and at
 the harbormaster pier at York Harbor in York, ME in May 2020. Colonies
 were transported to UNH in unfiltered seawater immediately after
 collection and were cultured and maintained as described above (Mackie,
 1966). To identify which texture to use in the larval settlement study, we
 tested three different textures along with a non-textured petri dish
 (smooth/unscratched). The experimental design of the pilot study was a 2x4
 split-plot RCBD with 7 blocks where the blocks were light boxes that
 contained two chambers. The main plot, which was the light condition, was
 applied on the chamber level and contained two levels: low green light
 (520nm; 5 µmol m-2 s-1) and darkness. The sub-plot was the dish texture
 created from scratches with different levels of sandpaper and contained a
 total of 4 levels: smooth (no scratches), fine (220-grit), medium
 (80-grit), and course (36-grit) (Lowes; model #s respectively:
 21220-SBP-15, 21080-SBP-15, 9150-052). The scratches were performed as
 previously mentioned, in a circular motion then with three scratches
 across, non-overlapping (right side, middle, and left side). The
 settlement study was performed as previously discussed with the same
 equipment where metamorphic stages were recorded at the following time
 points: 0 hour (hr), 2hr, 4hr, 6hr, 8hr, 12hr, and 24hr. The AUC values
 were calculated with the same equation from the settlement percentages as
 previously discussed and was analyzed using ANOVA and Tukey Rankings in
 the R environment.    Target sequences for RNA
 Fluorescent in situ hybridization probes Sequences
 were identified to make RNA probes for in situ hybridizations using the
 transcriptomic expression data from the developmental transcriptome study.
 We identified the highest expressed transcripts of key sensory genes from
 the three sensory gene sets. We then designed RNA-probes for our target
 sequences using the Stellaris RNA FISH platform (Biosearch Technologies)
 with the custom probe design service following their recommendations for
 probe design.
description: Microsoft Excel. Please see the README document (README.md). This
 dataset includes additional supplementary data for the article: Birch S.,
 and Plachetzki D. 2023. Multisensory integration by polymodal sensory
 neurons dictates settlement in actinulae larvae.