Immunohistochemical Approach to Understanding the Organization of the Olfactory System in the Cuttlefish, Sepia officinalis

Abstract

Cephalopods are nontraditional but captivating models of invertebrate neurobiology, particularly in evolutionary comparisons. Cephalopod olfactory systems have striking similarities and fundamental differences with vertebrates, arthropods, and gastropods, raising questions about the ancestral origins of those systems. We describe here the organization and development of the olfactory system of the common cuttlefish, Sepia officinalis, using immunohistochemistry and in situ hybridization. FMRFamide and/or related peptides and histamine are putative neurotransmitters in olfactory sensory neurons. Other neurotransmitters, including serotonin and APGWamide within the olfactory and other brain lobes, suggest efferent control of olfactory input and/or roles in the processing of olfactory information. The distributions of neurotransmitters, along with staining patterns of phalloidin, anti-acetylated α-tubulin, and a synaptotagmin riboprobe, help to clarify the structure of the olfactory lobe. We discuss a key difference, the lack of identifiable olfactory glomeruli, in cuttlefish in comparison to other models, and suggest its implications for the evolution of olfaction.

Histamine and histidine decarboxylase in the olfactory system and brain of the common cuttlefish Sepia officinalis (Linnaeus, 1758)

“…HA‐LIR has been observed previously in the olfactory lobes (Scaros et al, ) but reported difficulties in the penetration of the requisite fixatives prevented the extensive exploration of HA distribution using immunochemistry only. Therefore, we report our findings using HDC ISH and anti‐HA IHC when available.…”

“…Scaros et al () previously showed evidence of HA‐LIR in cells of the olfactory lobes. Using HDC ISH, we identified the presence of HDC‐expressing cells as well.…”

“…We were able to describe in additional detail the higher abundance of HDC cells in lobule 1 in comparison to lobules 2 and 3 at a higher resolution than was previously possible with IHC (Figure ). Scaros et al () also described HAergic fibers extending from the ventral end of the olfactory lobe that appears to form the olfactory nerve. In the same location, we observed a similar bundle of fibers extending from the ventral end of the olfactory lobe with HDC ISH (Figure c3).…”

“…All specimens used for experimentation were euthanized in isotonic magnesium chloride (7% in distilled water; dH 2 0) for 20 to 40 min until they became unresponsive to stimuli (Scaros et al, ). After euthanasia, embryos collected for immunohistochemistry (IHC) and ISH experiments were decapitated and the head region was prepared for fixation (ISH, n = 6; IHC, n = 11).…”

“…The two antibodies employed in this study were obtained commercially and were previously used in Scaros et al (). The HA antibody (Immunostar Cat# 22939, RRID: AB_582245) has been extensively used in vertebrates ( Mus ; McCann, Hwang, Hennig, Ward, & Sanders, ), arthropods ( Drosophila ; Stenesen, Moehlman, & Krämer, ), and molluscs ( Lymnaea , Hegedűs et al, ; Biomphalaria , Habib et al, ).…”