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Nitric Oxide Synthase in the Central Nervous System and Peripheral Organs of Stramonita haemastoma: Protein Distribution and Gene Expression in Response to Thermal Stress.

Toni M, De Angelis F, di Patti MC, Cioni C - Mar Drugs (2015)

Bottom Line: The detailed study of NOS distribution in peripheral and central neurons suggested that NOS is both intracellular and presynaptically located.Present findings confirm that NO may have a key role in the central neuronal circuits of gastropods and in sensory perception.The physiological relevance of NOS enzymes in the same organs was suggested by thermal stress experiments demonstrating that the constitutive expression of ShNOS is modulated in a time- and organ-dependent manner in response to environmental stressors.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology and Biotechnology "Charles Darwin", Sapienza University, 00161 Rome, Italy. mattia.toni@uniroma1.it.

ABSTRACT
Nitric oxide (NO) is generated via the oxidation of l-arginine by the enzyme NO synthase (NOS) both in vertebrates and invertebrates. Three NOS isoforms, nNOS, iNOS and eNOS, are known in vertebrates, whereas a single NOS isoform is usually expressed in invertebrates, sharing structural and functional characteristics with nNOS or iNOS depending on the species. The present paper is focused on the constitutive Ca(2+)/calmodulin-dependent nNOS recently sequenced by our group in the neogastropod Stramonita haemastoma (ShNOS). In this paper we provide new data on cellular distribution of ShNOS in the CNS (pedal ganglion) and peripheral organs (osphradium, tentacle, eye and foot) obtained by WB, IF, CM and NADPHd. Results demonstrated that NOS-like proteins are widely expressed in sensory receptor elements, neurons and epithelial cells. The detailed study of NOS distribution in peripheral and central neurons suggested that NOS is both intracellular and presynaptically located. Present findings confirm that NO may have a key role in the central neuronal circuits of gastropods and in sensory perception. The physiological relevance of NOS enzymes in the same organs was suggested by thermal stress experiments demonstrating that the constitutive expression of ShNOS is modulated in a time- and organ-dependent manner in response to environmental stressors.

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Osphradium histology. (A) Stereo microscope image of the osphradium that shows the bipinnate structure characterized by lamellae (l) radiating from the central axis (ca); (B) Frontal section of the osphradium showing the different cell density in the organ; (C) Sagittal section showing the lamellar axis (la) containing the lamellar nerve that sends a branch down (asterisks) into digitiform processes; (D) Frontal section of a single lamella showing its proximal-distal polarization characterized by sensory (sr), ciliated (cr) and glandular (gr) regions. Asterisks indicate the enlarged region in the lamellar axis (la) corresponding to the lamellar nerve branch observed in C; (E) Frontal section at the base of the lamella showing the lack of cell nuclei in the central axis and clusters of ganglionic cells (gc) at the base of the lamella; (F) Multilayered epithelium of the sensory region where S, Sz, Si3, Sch and am cells may be tentatively identified on the base of their morphology and position. The arrowhead indicates cilia on the apical portion of Sz cells. The arrow points to cell products secreted on the lamellar surface. Thin fibers running toward the epithelium surface can be glimpsed (double arrowheads); (G) Detail of the sensory epithelium in which yellowish granules in apical cytoplasm of S cells can be observed. Note the absence of these granules in Sz cells; (H) Apical process (double arrowhead) of a sensory Si3 cell (arrowhead) contacting the epithelium surface is shown; (I) Frontal section showing the lamellar groove (grv) that separates the sensory and ciliate regions. In the distal portion of the sensory region, Si4 cells can be observed. Sensory dendrites are regularly intercalated to epithelial cells on the apical surface of the epithelium (double arrowheads). Arrowhead points to Shc mucous cells. At the base of the groove, putative Si1 cells can be identified. Arrow points to the cilia of Si2 cells in the ciliated region; (J) Detail showing a Si4 cell with its shape at “halved onion”; (K) Image of neighboring ciliated and glandular regions showing the high concentration of gland cells in the latter; (L) Detail of the glandular region in which two different types of gland cells can be distinguished on the base of their hematoxylin staining: well-stained gland cells (am) and cells with low affinity for hematoxylin (glc). The nuclei are visible at the cell base (arrowhead). Gland cells secrete their products in the interlamellar space (ils). Among glands, support epithelial cells are present (arrow); (M) Frontal section of the lamellar tip where a different cell distribution can be observed. Thin fibers coursing from the lamellar axis to the epithelium surface are observed among epithelial cells (arrows). Bars: A = 500 μm; B = 150 μm; C = 100 μm; D, E = 50 μm; F, G, I–L = 10 μm; H = 5 μm; M = 25 μm.
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marinedrugs-13-06636-f002: Osphradium histology. (A) Stereo microscope image of the osphradium that shows the bipinnate structure characterized by lamellae (l) radiating from the central axis (ca); (B) Frontal section of the osphradium showing the different cell density in the organ; (C) Sagittal section showing the lamellar axis (la) containing the lamellar nerve that sends a branch down (asterisks) into digitiform processes; (D) Frontal section of a single lamella showing its proximal-distal polarization characterized by sensory (sr), ciliated (cr) and glandular (gr) regions. Asterisks indicate the enlarged region in the lamellar axis (la) corresponding to the lamellar nerve branch observed in C; (E) Frontal section at the base of the lamella showing the lack of cell nuclei in the central axis and clusters of ganglionic cells (gc) at the base of the lamella; (F) Multilayered epithelium of the sensory region where S, Sz, Si3, Sch and am cells may be tentatively identified on the base of their morphology and position. The arrowhead indicates cilia on the apical portion of Sz cells. The arrow points to cell products secreted on the lamellar surface. Thin fibers running toward the epithelium surface can be glimpsed (double arrowheads); (G) Detail of the sensory epithelium in which yellowish granules in apical cytoplasm of S cells can be observed. Note the absence of these granules in Sz cells; (H) Apical process (double arrowhead) of a sensory Si3 cell (arrowhead) contacting the epithelium surface is shown; (I) Frontal section showing the lamellar groove (grv) that separates the sensory and ciliate regions. In the distal portion of the sensory region, Si4 cells can be observed. Sensory dendrites are regularly intercalated to epithelial cells on the apical surface of the epithelium (double arrowheads). Arrowhead points to Shc mucous cells. At the base of the groove, putative Si1 cells can be identified. Arrow points to the cilia of Si2 cells in the ciliated region; (J) Detail showing a Si4 cell with its shape at “halved onion”; (K) Image of neighboring ciliated and glandular regions showing the high concentration of gland cells in the latter; (L) Detail of the glandular region in which two different types of gland cells can be distinguished on the base of their hematoxylin staining: well-stained gland cells (am) and cells with low affinity for hematoxylin (glc). The nuclei are visible at the cell base (arrowhead). Gland cells secrete their products in the interlamellar space (ils). Among glands, support epithelial cells are present (arrow); (M) Frontal section of the lamellar tip where a different cell distribution can be observed. Thin fibers coursing from the lamellar axis to the epithelium surface are observed among epithelial cells (arrows). Bars: A = 500 μm; B = 150 μm; C = 100 μm; D, E = 50 μm; F, G, I–L = 10 μm; H = 5 μm; M = 25 μm.

Mentions: As in other mollusks, the osphradium of S. haemastoma is a bipinnate organ consisting of two opposite rows of flat epithelial lamellae radiating from the central axis (Figure 2A–C). The osphradium contains the osphradial ganglion which has an inner neuropile surrounded by ganglionar cell bodies (Figure 2B). Ganglion cells are clustered in correspondence to the interlamellar spaces (Figure 2B,E). Osphradial lamellae radiate from the central axis and each lamella receives one branch of the lamellar nerve which forms its longitudinal axis (Figure 2C,D). Among lamellar nerve fibers, small neurons were aligned in the lamellar axis (Figure 2D).


Nitric Oxide Synthase in the Central Nervous System and Peripheral Organs of Stramonita haemastoma: Protein Distribution and Gene Expression in Response to Thermal Stress.

Toni M, De Angelis F, di Patti MC, Cioni C - Mar Drugs (2015)

Osphradium histology. (A) Stereo microscope image of the osphradium that shows the bipinnate structure characterized by lamellae (l) radiating from the central axis (ca); (B) Frontal section of the osphradium showing the different cell density in the organ; (C) Sagittal section showing the lamellar axis (la) containing the lamellar nerve that sends a branch down (asterisks) into digitiform processes; (D) Frontal section of a single lamella showing its proximal-distal polarization characterized by sensory (sr), ciliated (cr) and glandular (gr) regions. Asterisks indicate the enlarged region in the lamellar axis (la) corresponding to the lamellar nerve branch observed in C; (E) Frontal section at the base of the lamella showing the lack of cell nuclei in the central axis and clusters of ganglionic cells (gc) at the base of the lamella; (F) Multilayered epithelium of the sensory region where S, Sz, Si3, Sch and am cells may be tentatively identified on the base of their morphology and position. The arrowhead indicates cilia on the apical portion of Sz cells. The arrow points to cell products secreted on the lamellar surface. Thin fibers running toward the epithelium surface can be glimpsed (double arrowheads); (G) Detail of the sensory epithelium in which yellowish granules in apical cytoplasm of S cells can be observed. Note the absence of these granules in Sz cells; (H) Apical process (double arrowhead) of a sensory Si3 cell (arrowhead) contacting the epithelium surface is shown; (I) Frontal section showing the lamellar groove (grv) that separates the sensory and ciliate regions. In the distal portion of the sensory region, Si4 cells can be observed. Sensory dendrites are regularly intercalated to epithelial cells on the apical surface of the epithelium (double arrowheads). Arrowhead points to Shc mucous cells. At the base of the groove, putative Si1 cells can be identified. Arrow points to the cilia of Si2 cells in the ciliated region; (J) Detail showing a Si4 cell with its shape at “halved onion”; (K) Image of neighboring ciliated and glandular regions showing the high concentration of gland cells in the latter; (L) Detail of the glandular region in which two different types of gland cells can be distinguished on the base of their hematoxylin staining: well-stained gland cells (am) and cells with low affinity for hematoxylin (glc). The nuclei are visible at the cell base (arrowhead). Gland cells secrete their products in the interlamellar space (ils). Among glands, support epithelial cells are present (arrow); (M) Frontal section of the lamellar tip where a different cell distribution can be observed. Thin fibers coursing from the lamellar axis to the epithelium surface are observed among epithelial cells (arrows). Bars: A = 500 μm; B = 150 μm; C = 100 μm; D, E = 50 μm; F, G, I–L = 10 μm; H = 5 μm; M = 25 μm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4663546&req=5

marinedrugs-13-06636-f002: Osphradium histology. (A) Stereo microscope image of the osphradium that shows the bipinnate structure characterized by lamellae (l) radiating from the central axis (ca); (B) Frontal section of the osphradium showing the different cell density in the organ; (C) Sagittal section showing the lamellar axis (la) containing the lamellar nerve that sends a branch down (asterisks) into digitiform processes; (D) Frontal section of a single lamella showing its proximal-distal polarization characterized by sensory (sr), ciliated (cr) and glandular (gr) regions. Asterisks indicate the enlarged region in the lamellar axis (la) corresponding to the lamellar nerve branch observed in C; (E) Frontal section at the base of the lamella showing the lack of cell nuclei in the central axis and clusters of ganglionic cells (gc) at the base of the lamella; (F) Multilayered epithelium of the sensory region where S, Sz, Si3, Sch and am cells may be tentatively identified on the base of their morphology and position. The arrowhead indicates cilia on the apical portion of Sz cells. The arrow points to cell products secreted on the lamellar surface. Thin fibers running toward the epithelium surface can be glimpsed (double arrowheads); (G) Detail of the sensory epithelium in which yellowish granules in apical cytoplasm of S cells can be observed. Note the absence of these granules in Sz cells; (H) Apical process (double arrowhead) of a sensory Si3 cell (arrowhead) contacting the epithelium surface is shown; (I) Frontal section showing the lamellar groove (grv) that separates the sensory and ciliate regions. In the distal portion of the sensory region, Si4 cells can be observed. Sensory dendrites are regularly intercalated to epithelial cells on the apical surface of the epithelium (double arrowheads). Arrowhead points to Shc mucous cells. At the base of the groove, putative Si1 cells can be identified. Arrow points to the cilia of Si2 cells in the ciliated region; (J) Detail showing a Si4 cell with its shape at “halved onion”; (K) Image of neighboring ciliated and glandular regions showing the high concentration of gland cells in the latter; (L) Detail of the glandular region in which two different types of gland cells can be distinguished on the base of their hematoxylin staining: well-stained gland cells (am) and cells with low affinity for hematoxylin (glc). The nuclei are visible at the cell base (arrowhead). Gland cells secrete their products in the interlamellar space (ils). Among glands, support epithelial cells are present (arrow); (M) Frontal section of the lamellar tip where a different cell distribution can be observed. Thin fibers coursing from the lamellar axis to the epithelium surface are observed among epithelial cells (arrows). Bars: A = 500 μm; B = 150 μm; C = 100 μm; D, E = 50 μm; F, G, I–L = 10 μm; H = 5 μm; M = 25 μm.
Mentions: As in other mollusks, the osphradium of S. haemastoma is a bipinnate organ consisting of two opposite rows of flat epithelial lamellae radiating from the central axis (Figure 2A–C). The osphradium contains the osphradial ganglion which has an inner neuropile surrounded by ganglionar cell bodies (Figure 2B). Ganglion cells are clustered in correspondence to the interlamellar spaces (Figure 2B,E). Osphradial lamellae radiate from the central axis and each lamella receives one branch of the lamellar nerve which forms its longitudinal axis (Figure 2C,D). Among lamellar nerve fibers, small neurons were aligned in the lamellar axis (Figure 2D).

Bottom Line: The detailed study of NOS distribution in peripheral and central neurons suggested that NOS is both intracellular and presynaptically located.Present findings confirm that NO may have a key role in the central neuronal circuits of gastropods and in sensory perception.The physiological relevance of NOS enzymes in the same organs was suggested by thermal stress experiments demonstrating that the constitutive expression of ShNOS is modulated in a time- and organ-dependent manner in response to environmental stressors.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology and Biotechnology "Charles Darwin", Sapienza University, 00161 Rome, Italy. mattia.toni@uniroma1.it.

ABSTRACT
Nitric oxide (NO) is generated via the oxidation of l-arginine by the enzyme NO synthase (NOS) both in vertebrates and invertebrates. Three NOS isoforms, nNOS, iNOS and eNOS, are known in vertebrates, whereas a single NOS isoform is usually expressed in invertebrates, sharing structural and functional characteristics with nNOS or iNOS depending on the species. The present paper is focused on the constitutive Ca(2+)/calmodulin-dependent nNOS recently sequenced by our group in the neogastropod Stramonita haemastoma (ShNOS). In this paper we provide new data on cellular distribution of ShNOS in the CNS (pedal ganglion) and peripheral organs (osphradium, tentacle, eye and foot) obtained by WB, IF, CM and NADPHd. Results demonstrated that NOS-like proteins are widely expressed in sensory receptor elements, neurons and epithelial cells. The detailed study of NOS distribution in peripheral and central neurons suggested that NOS is both intracellular and presynaptically located. Present findings confirm that NO may have a key role in the central neuronal circuits of gastropods and in sensory perception. The physiological relevance of NOS enzymes in the same organs was suggested by thermal stress experiments demonstrating that the constitutive expression of ShNOS is modulated in a time- and organ-dependent manner in response to environmental stressors.

Show MeSH
Related in: MedlinePlus