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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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Tentacle: histology and NADPHd staining. (A,B) Lateral and frontal view of the tentacle observed at the stereo microscope. The arrow points to the eye; (C) Light microscopy of a sagittal section of the tentacle (stained by hematoxylin/eosin) showing the eye (arrow) and the folded surface (arrowheads); (D) Detail of the folded surface covered by a ciliated columnar epithelium; (E) Magnification of the epithelium showing the presence of cilia (arrows). Brownish granules are seen on the apical portion of the cells (double arrowhead). Dark gland cells are evident among epithelial elements (arrowheads); (F) Gland cells release their secretion on the tentacle surface (arrowheads); (G) Microphotograph of the eye where the basal retinal neurons (BRN) and the pigmented region (PR) are visible; (H) Sagittal section showing the optic nerve (ON); (I) Detail of RL contains distal segments of photoreceptors, PR contains retinal pigmented cells and BRN is formed by retinal neurons; (J–L) NADPHd assay on sagittal sections of the foot; (J) Microphotograph of the eye showing NADPHd blue staining in the optic nerve (ON), and the BRN layer. Intense NADPHd staining is present in the external BRN layer. The dark pigmentation of the PR prevents the detection of NADPHd staining; (K) Weak and diffuse NADPHd staining of epithelial cells. Apical pigments obscure NADPHd staining at the cell apex; (L) NADPHd-positive gland cells are seen within the epithelium (arrow). Bars: A–C = 500 μm; D = 50 μm; E, G, H = 10 μm; I = 5 μm; F, J–K = 25 μm.
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marinedrugs-13-06636-f004: Tentacle: histology and NADPHd staining. (A,B) Lateral and frontal view of the tentacle observed at the stereo microscope. The arrow points to the eye; (C) Light microscopy of a sagittal section of the tentacle (stained by hematoxylin/eosin) showing the eye (arrow) and the folded surface (arrowheads); (D) Detail of the folded surface covered by a ciliated columnar epithelium; (E) Magnification of the epithelium showing the presence of cilia (arrows). Brownish granules are seen on the apical portion of the cells (double arrowhead). Dark gland cells are evident among epithelial elements (arrowheads); (F) Gland cells release their secretion on the tentacle surface (arrowheads); (G) Microphotograph of the eye where the basal retinal neurons (BRN) and the pigmented region (PR) are visible; (H) Sagittal section showing the optic nerve (ON); (I) Detail of RL contains distal segments of photoreceptors, PR contains retinal pigmented cells and BRN is formed by retinal neurons; (J–L) NADPHd assay on sagittal sections of the foot; (J) Microphotograph of the eye showing NADPHd blue staining in the optic nerve (ON), and the BRN layer. Intense NADPHd staining is present in the external BRN layer. The dark pigmentation of the PR prevents the detection of NADPHd staining; (K) Weak and diffuse NADPHd staining of epithelial cells. Apical pigments obscure NADPHd staining at the cell apex; (L) NADPHd-positive gland cells are seen within the epithelium (arrow). Bars: A–C = 500 μm; D = 50 μm; E, G, H = 10 μm; I = 5 μm; F, J–K = 25 μm.

Mentions: The two eyes of S. haemastoma are bilaterally placed on the eyestalks, fused to the posterior edge of the cephalic tentacles. In stereo microscopy, tentacles appear dark brown because of the pigments contained in the epithelium (Figure 4A–C). Each tentacle is covered by a ciliated columnar epithelium provided with brown pigments and is rich in gland cells (Figure 4D–F). The presence of pigments, including carotenoids, indigoids, melanin, porphyrins and bilichromes, is a common feature in mollusk epithelia.


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)

Tentacle: histology and NADPHd staining. (A,B) Lateral and frontal view of the tentacle observed at the stereo microscope. The arrow points to the eye; (C) Light microscopy of a sagittal section of the tentacle (stained by hematoxylin/eosin) showing the eye (arrow) and the folded surface (arrowheads); (D) Detail of the folded surface covered by a ciliated columnar epithelium; (E) Magnification of the epithelium showing the presence of cilia (arrows). Brownish granules are seen on the apical portion of the cells (double arrowhead). Dark gland cells are evident among epithelial elements (arrowheads); (F) Gland cells release their secretion on the tentacle surface (arrowheads); (G) Microphotograph of the eye where the basal retinal neurons (BRN) and the pigmented region (PR) are visible; (H) Sagittal section showing the optic nerve (ON); (I) Detail of RL contains distal segments of photoreceptors, PR contains retinal pigmented cells and BRN is formed by retinal neurons; (J–L) NADPHd assay on sagittal sections of the foot; (J) Microphotograph of the eye showing NADPHd blue staining in the optic nerve (ON), and the BRN layer. Intense NADPHd staining is present in the external BRN layer. The dark pigmentation of the PR prevents the detection of NADPHd staining; (K) Weak and diffuse NADPHd staining of epithelial cells. Apical pigments obscure NADPHd staining at the cell apex; (L) NADPHd-positive gland cells are seen within the epithelium (arrow). Bars: A–C = 500 μm; D = 50 μm; E, G, H = 10 μm; I = 5 μm; F, J–K = 25 μm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4663546&req=5

marinedrugs-13-06636-f004: Tentacle: histology and NADPHd staining. (A,B) Lateral and frontal view of the tentacle observed at the stereo microscope. The arrow points to the eye; (C) Light microscopy of a sagittal section of the tentacle (stained by hematoxylin/eosin) showing the eye (arrow) and the folded surface (arrowheads); (D) Detail of the folded surface covered by a ciliated columnar epithelium; (E) Magnification of the epithelium showing the presence of cilia (arrows). Brownish granules are seen on the apical portion of the cells (double arrowhead). Dark gland cells are evident among epithelial elements (arrowheads); (F) Gland cells release their secretion on the tentacle surface (arrowheads); (G) Microphotograph of the eye where the basal retinal neurons (BRN) and the pigmented region (PR) are visible; (H) Sagittal section showing the optic nerve (ON); (I) Detail of RL contains distal segments of photoreceptors, PR contains retinal pigmented cells and BRN is formed by retinal neurons; (J–L) NADPHd assay on sagittal sections of the foot; (J) Microphotograph of the eye showing NADPHd blue staining in the optic nerve (ON), and the BRN layer. Intense NADPHd staining is present in the external BRN layer. The dark pigmentation of the PR prevents the detection of NADPHd staining; (K) Weak and diffuse NADPHd staining of epithelial cells. Apical pigments obscure NADPHd staining at the cell apex; (L) NADPHd-positive gland cells are seen within the epithelium (arrow). Bars: A–C = 500 μm; D = 50 μm; E, G, H = 10 μm; I = 5 μm; F, J–K = 25 μm.
Mentions: The two eyes of S. haemastoma are bilaterally placed on the eyestalks, fused to the posterior edge of the cephalic tentacles. In stereo microscopy, tentacles appear dark brown because of the pigments contained in the epithelium (Figure 4A–C). Each tentacle is covered by a ciliated columnar epithelium provided with brown pigments and is rich in gland cells (Figure 4D–F). The presence of pigments, including carotenoids, indigoids, melanin, porphyrins and bilichromes, is a common feature in mollusk epithelia.

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