Glial cells in the posterior sub-esophageal mass of the brain in Sepia officinalis (Linnaeus, 1758) (decapodiformes-sepiida): ultrastructure and cytochemical studies.

Electron microscopy revealed that glial cells in the posterior sub-esophageal mass of the brain in Sepia officinalis had a well-developed rough endoplasmic reticulum formed by long coverslips with rectilinear or curvilinear arrangements. The coverslips appeared dilated and have a large amount of adhered polysomes. Vesicular lamellae coexisted with the elongated lamellae of RER and dictyosomes of Golgi apparatus. Endocytosis was evidenced through the pale vesicles which were appeared next to the apical border of microvilli in some glial cells. Sub-cellular features of endocytosis, predominantly the fluid phase, were observed in the apical glial cell cytoplasm. Glial cells were related to phagocytosis of apoptotic neurons, endocytosis, pinocytosis and adsorption. These functions were proposed based on their ultrastructure characteristics and a significant number of vesicles with different shapes (oval to polygonal), sizes 0.052-0.67 µm and contents. Glycogen, MPS and lipid were detected in the glial cells. Alkaline phosphatase was not observed, while an activity of acid phosphatase was bound to lysosomes. ATPases were present in the glial cells along the lateral and basal plasma lemma as well as on the membranes of cell organelles. Unspecific esterase was clearly recognizable by electron microscopy. The monoamine and cytochrome oxidase activities were demonstrated, while the succinate dehydrogenase showed a weak enzyme activity.

Morphology of reproductive accessory glands in female Sepia pharaonis (Cephalopoda: Sepiidae) sheds light on egg encapsulation.

The pharaoh cuttlefish, Sepia pharaonis, is an important cephalopod fishery species in southeastern Asia, with understudied reproductive physiology. The present study aimed to investigate the cellular characteristics of epithelial cells found in the nidamental glands (NGs) and accessory NGs (ANGs), as well as the structural connections between these two glands in mature female S. pharaonis. A histological analysis revealed two types of epithelial cells in NGs: Alcian blue-positive, PAS-negative mucosubstance-secreting cells and eosinophilic, PAS-positive granule-secreting cells. Using transmission electron microscopy, three types of epithelial cells were identified: cells with electron-dense granules, cells with electron-lucent granules, and cells with both cilia and microvilli in the apex. Mature ANGs contain an abundance of tubular units composed of epithelial cells resting on a thin layer of basal lamina. Innervated muscle cells are tightly adhered to the basal lamina. In addition, we observed epithelial canalization of ANG tubules penetrating through the connective tissue linking NGs and the walls of the tubules in ANGs, which allows the contents of the ANG tubules to be transported to the NGs. Our results suggest that ANGs participate in the encapsulation of the ova via the same pathway as NGs, which provides an important basis for future studies on the mechanism of protection provided by NGs and ANGs during embryonic development in S. pharaonis.

Microanatomy and ultrastructure of outer mantle epidermis of the cuttlefish, Sepia esculenta (Cephalopoda: Sepiidae).

This study describes the ultrastructural characteristics of external epidermis of mantle of Sepia esculenta using light and electron microscopy. The epidermis was thicker on the ventral surface than on the dorsal surface, with a higher secretory cell distribution on the ventral surface than on the dorsal surface. The epidermis was a single layer composed of epithelial cells, secretory cells, ciliated cells and neuroglial cells. Epithelial cells were columnar with well-developed microvilli on the free surface, and the microvilli were covered with glycocalyx. The epithelial cells were connected to the neighboring cells by tight junctions and membrane interdigitations of the apico-frontal surface. Well-developed microfilaments were arranged in a vertical direction in the cortical cytoplasm. The secretory cells were categorized into three types (A, B and C) in accordance with the light microscopical characteristics and ultrastructures of the secretory granules. The distribution of these cells was in the following order: Type A>Type B>Type C. SEM observation revealed that the secretory pore size of the Type A secretory cells was approximately 8.6 µm×12.2 µm. Cytoplasm displayed a red color as the result of Masson's trichrome stain and H-E stain, and contained polygonal granules of approximately 1.2 µm2 with a high electron density. The secretory pore size of the Type B secretory cells was approximately 10.1 µm×12.1 µm. As the results of AB-PAS (pH 2.5) and AF-AB (pH 2.5) reactions, the cytoplasm displayed a red color. The cells contained membrane bounded secretory granules with very low electron density. The secretory pore of the Type C secretory cells was circular shape, and approximately 5.5 µm×5.5 µm. Cytoplasm was found to be homogeneous under H-E stain and Masson's trichrome stain, and displayed a red color. As the result of AB-PAS (pH 2.5) reaction, the cytoplasm displayed a red color. The electron density of the secretory substance was the highest among the three types of secretory cells. The ciliated cells had a ciliary tuft on the free surface and were distributed throughout the mantle with the exception of the adhesive organs. Neuroglial cells were connected to the basal membrane, epithelial cells, secretory cells and nerve fibers through cytoplasmic process, and contained neurosecretory granules with high electron density within the cytoplasm.

Ancient origin of somatic and visceral neurons.

BACKGROUND: A key to understanding the evolution of the nervous system on a large phylogenetic scale is the identification of homologous neuronal types. Here, we focus this search on the sensory and motor neurons of bilaterians, exploiting their well-defined molecular signatures in vertebrates. Sensorimotor circuits in vertebrates are of two types: somatic (that sense the environment and respond by shaping bodily motions) and visceral (that sense the interior milieu and respond by regulating vital functions). These circuits differ by a small set of largely dedicated transcriptional determinants: Brn3 is expressed in many somatic sensory neurons, first and second order (among which mechanoreceptors are uniquely marked by the Brn3+/Islet1+/Drgx+ signature), somatic motoneurons uniquely co-express Lhx3/4 and Mnx1, while the vast majority of neurons, sensory and motor, involved in respiration, blood circulation or digestion are molecularly defined by their expression and dependence on the pan-visceral determinant Phox2b. RESULTS: We explore the status of the sensorimotor transcriptional code of vertebrates in mollusks, a lophotrochozoa clade that provides a rich repertoire of physiologically identified neurons. In the gastropods Lymnaea stagnalis and Aplysia californica, we show that homologues of Brn3, Drgx, Islet1, Mnx1, Lhx3/4 and Phox2b differentially mark neurons with mechanoreceptive, locomotory and cardiorespiratory functions. Moreover, in the cephalopod Sepia officinalis, we show that Phox2 marks the stellate ganglion (in line with the respiratory--that is, visceral--ancestral role of the mantle, its target organ), while the anterior pedal ganglion, which controls the prehensile and locomotory arms, expresses Mnx. CONCLUSIONS: Despite considerable divergence in overall neural architecture, a molecular underpinning for the functional allocation of neurons to interactions with the environment or to homeostasis was inherited from the urbilaterian ancestor by contemporary protostomes and deuterostomes.

Principles underlying chromatophore addition during maturation in the European cuttlefish, Sepia officinalis.

The goal of this work was to identify some of the principles underlying chromatophore growth and development in the European cuttlefish, Sepia officinalis. One set of experiments used a regeneration model to follow the re-growth of black chromatophores for 30 days following excision of a small piece of fin tissue. A separate set of experiments tracked and analyzed the addition of new fin chromatophores during a month of normal, undisturbed growth. We also followed the development of individual chromatophores from their initial appearance to full maturation to determine whether their color type was fixed. Based on the results of these studies, we propose five guiding principles for chromatophore growth and maturation. (1) The three chromatophore cell types--black, reddish-brown and yellow--are present at different spatial frequencies in the cuttlefish fin. (2) During normal growth, new chromatophores are inserted at a higher spatial frequency than existing (control) chromatophores of the same color type. (3) In regenerating tissue, new black chromatophores are initially added at low spatial frequencies. As regeneration continues, new black chromatophores appear at increasing spatial frequencies until they are inserted at a spatial frequency higher than that observed in control tissue, similar to the way in which chromatophores were observed to be added in normally growing tissue. (4) All chromatophores first appear as pale orange cells and slowly darken into their respective color types without passing through intermediate color stages. (5) New black chromatophores undergo a doubling in size as they mature, while reddish-brown and yellow chromatophores do not grow at all after they are inserted in the dermis.

Evidence of early nervous differentiation and early catecholaminergic sensory system during Sepia officinalis embryogenesis.

Within Mollusca, cephalopods exhibit a particularly complex nervous system. The adult brain is formed from the fusion of several "typical" molluscan ganglia but it remains poorly understood how these ganglia emerge, migrate, and differentiate during embryogenesis. We studied the development of both central and peripheral nervous system by antibodies raised against alpha-tubulin and tyrosine hydroxylase (TH) in Sepia officinalis embryos to visualize neurites and catecholamine-containing neurons, respectively. In early embryos, when organs start delineating, some ganglia already exhibited a significant fiber network. TH-like immunoreactivity was detected in these fibers and in some primary sensory neurons in the embryo periphery. These data attest to the occurrence of an early embryonic sensory nervous system, likely effective, transient in part, and in relation to the perception of external cues. Concerning the peripheral nervous network, the stellate ganglia emerged as a plexus of numerous converging axons from TH-like immunoreactive sensory cells, first at the mantle edge, and then in the whole mantle surface. Later, TH-immunopositive motor fibers, originating from the stellate ganglia, penetrated the circular muscles of the mantle. These patterns reveal the setup of a mantle midline with likely attractive and repulsive properties. Our findings seem to challenge the widespread, still accepted, view of a late differentiation of cephalopod ganglia, and provides significant data for further investigations about axonal guidance during cephalopod development.

Sperm nucleomorphogenesis in the cephalopod Sepia officinalis.

Sperm nucleomorphogenesis in the cephalopod Sepia officinalis is the product of the interaction between perinuclear microtubules and condensing chromatin. This interaction occurs during spermiogenesis and is established through the nuclear membrane. As in other cephalopod species, the perinuclear microtubules are transient structures. In the case of S. officinalis, they begin to appear in the basal area of the early spermatid and progress from there, establishing contact with the external nuclear membrane and follow a defined, but not symmetric, geometry. Thus, the microtubules accumulate preferentially in one area of the nuclear membrane which we refer to here as the "dorsal zone". Later, the microtubules will be eliminated before the mature spermatid migrates to the epidydimis. The chromatin is condensed within the nucleus following a complex pattern, beginning as fibro-granular structures until forming fibres of approximately 45 nm diameter (patterning phases). From this stage on, an increase in the chemical basicity of DNA-interacting proteins is produced, and chromatin fibres coalesce together, being recruited to the dorsal zone of the membrane, where there is a higher density of microtubules. This last step (condensation phases) allows the chromatin fibres to be arranged parallel to the axis of the elongating nucleus, and more importantly, is deduced to cause a lateral compression of the nucleus. This lateral compression is in fact a recruitment of the ventral zone toward the dorsal zone, which brings about an important reduction in nuclear volume. The detailed observations which comprise this work complement previous studies of spermiogenesis of Sepia and other cephalopods, and will help to better understand the process of cellular morphology implicated in the evolution of sperm nuclear shape in this taxonomic group.