Comparative study of the sterol composition of the digestive gland and the gonad of Sepia officinalis L. (mollusca,cephalopoda)

• 1.1. The sterol composition of the digestive gland and the gonad of Sepia officinalis L. was investigated by GC and GC-MS.
• 2.2. The same sterols were recognized in both organs, cholesterol being the major component of the sterol mixtures. However, quantitative differences appeared between the sterol composition of the digestive gland and the gonad.
• 3.3. The sterol mixtures of the digestive gland and the gonad of immature and mature females and males of various origins were compared. Quantitative changes in the sterol composition of the gonad were related to sexual maturity whereas the sterol composition of the digestive gland appeared linked to the diet.

     

Evidence for a molecular polymorphis of cholinesterase in Sepia officinalis (cephalopoda: decapoda)

• 1.1. Three forms of cholinesterase were sequentially extracted from head and tentacles of Sepia officinalis and noted as low-salt (LSS), detergent (DS) and high-salt (HSS) soluble. They represent about 24, 30 and 46% of total activity.
• 2.2. All enzyme forms seem to be amphiphilic proteins with hydrophobic domains interacting with non-ionic detergent (Triton X-100) and giving self-aggregation (LSS form).
• 3.3. The DS form is membrane-anchored by a phosphatidylinositol, while the HSS form is likely linked to some proteoglycan molecule of the extracellular matrix by ionic interactions.
• 4.4. According to V_max/K_m values, all the enzymes are acetylcholinesterases, even if hydrolyze propionylthiocoline at the highest rate.
• 5.5. Some kinetic and molecular properties of the studied enzymes are compared with those of other cholinesterases from vertebrates and invertebrates. Possible phylogenic and adaptive features are discussed.

     

Cytomorphological aspects on the response of the branchial heart complex of Sepia officinalis L. (cephalopoda) to xenobiotics and bacterial infection

The ovoid cells of the branchial heart complex of Sepia officinalis L. were investigated with respect to their role in detoxification processes. The electron microscopical localization of in situ injected ferritin in the endocytotic-lysosomal system, and the fluorescence microscopical localization of protein-bound Evans blue in the ovoid cells of in vivo incubated animals, indicate that foreign materials are eliminated from the hemolymph by the branchial heart tissue. In addition to the non-circulating ovoid cells of the branchial heart, hemocytes in the circulating blood and in the wall of the branchial heart are also involved in the incorporation of allogeneic substances and bacteria, or their debris. Based on these observations, we propose that the ovoid cells, together with circulating and adhesive hemocytes in the branchial hearts, are an important component of a more comprehensive defence and detoxification system in dibranchiate cephalopods that prevents contamination of the whole organism by endocytotic removal of noxious substances from the hemolymph.     

Localization of catecholamine-containing nerve fibres in the branchial heart and cardiac ganglion of the common cuttlefish Sepia officinalis L. (cephalopoda)

The innervation of the branchial heart of Sepia officinalis was examined using TEM and glyoxylic acid induced fluorescence. In the cardiac ganglion and in cardiac nerves bluish-green fluorophores were seen associated with perikarya and varicose nerve fibres. Microspectrofluorometric analysis provided clear evidence that monoaminergic neurons in the branchial heart contain only catecholamines. Considering pharmacological data, it is more than likely that 5-hydroxytryptamine (serotonin) is not present in this system.     

Peptidergic control of egg-laying in the cephalopod Sepia officinalis: involvement of FMRFamide and FMRFamide-related peptides.

The peptidergic control of egg-laying was investigated in Sepia officinalis by using a myotropic bioassay. Three myotropic high-performance liquid chromatography fractions were obtained from optic lobe extracts. In the first fraction, FMRFamide (FMRFa) and FLRFa were isolated and sequenced. FMRFa-related peptides then were sought by dotting immunobinding of optic lobes extracts. The four immunoreactive fractions detected revealed the occurrence of FMRFa, FLRFa, FIRFa, and ALSGDAFLRFa predicted by the precursor already cloned from the optic lobes of S. officinalis (J Exp Biol 200:1483-9;1997). These peptides clearly appeared to be involved in the regulation of oocyte transport through the oviduct: the tetrapeptides FMRFa and FLRFa stimulated the contractions, whereas FIRFa and ALSGDAFLRFa lowered the tonus, the frequency, and the amplitude of the contractions. The occurrence of FaRPs in the nervous endings of the accessory sex glands suggested that this peptide family is involved in the regulation of secretory processes of the egg capsule. Indeed, FMRFa modulates the contractions of the main nidamental glands in vitro and, thus, should induce mechanical release of the secretion in vivo during ovulation. These results show that the FaRPs could play an important role in the synchronization of ovulation and egg capsule coating.     

The localization of enzyme activities in the pancreatic appendages of Sepia officinalis L. (Cephalopoda)

Summary In this study, enzyme activities of the pancreatic appendages of the ductus hepatopancreas (the so-called Pancreas) in Sepia officinalis L. have been demonstrated by light and electron microscopical methods: Malate dehydrogenase, monoamine oxidase, acid phosphatase, -glucuronidase, adenosine triphosphatase and carbonic anhydrase were shown by the former, and monoamine oxidase, catalase, glutamic oxalacetic transaminase, choline esterase (non-specific), alkaline phosphatase, acid phosphatase and carbonic anhydrase by the latter technique.The correlation between enzyme activity and distribution, and the presumed function of the two pancreatic epithelia is discussed.This study was supported by the Deutsche Forschungsgemeinschaft     

Adenosine triphosphatase localization in the branchial heart of Sepia officinalis L. (Cephalopoda)

Summary Sodium- and potassium-dependent adenosine triphosphatase (Na⁺–K⁺-ATPase) is demonstrated in the branchial heart of Sepia officinalis L. by biochemical, cytochemical and autoradiographical methods. The biochemical data indicate the presence of Na⁺–K⁺-ATPase, shown by potassium and magnesium dependency and inhibition by ouabain. Cytochemically and autoradiographically, the enzyme is localized in the sarcolemma of the muscle cells. The positive reaction of the transparent cells (type I cells) is due to activity of alkaline phosphatases. The dark cells (type II cells) react negatively. In addition to the Na⁺–K⁺-ATPase, a magnesium-activated adenosine triphosphatase (Mg²⁺-ATPase) and a bicarbonate-stimulated ATPase (HCO ₃ ^- -ATPase) are localized in the mitochondria.This study was supported by the Deutsche Forschungsgemeinschaft and is part of the doctoral dissertation     

Adenosine triphosphatase localization in the branchial heart of Sepia officinalis L. (Cephalopoda)

Sodium- and potassium-dependent adenosine triphosphatase (Na+--K+-ATPase) is demonstrated in the branchial heart of Sepia officinalis L. by biochemical, cytochemical and autoradiographical methods. The biochemical data indicate the presence of Na+--K+-ATPase, shown by potassium and magnesium dependency and inhibition by ouabain. Cytochemically and autoradiographically, the enzyme is localized in the sarcolemma of the muscle cells. The positive reaction of the transparent cells (type I cells) is due to activity of alkaline phosphatases. The dark cells (type II cells) react negatively. In addition to the Na+--K+-ATPase, a magnesium-activated adenosine triphosphatase (Mg2+-ATPase) and a bicarbonate-stimulated ATPase (HCO3(-)-ATPase) are localized in the mitochondria.     

Adenosine triphosphatase localization in the branchial heart appendage of Sepia officinalis L. (Cephalopoda)

Summary Sodium- and potassium-dependent adenosine triphosphatase (Na⁺–K⁺-ATPase) has been demonstrated in the branchial heart appendage (pericardial gland) of Sepia officinalis L. by biochemical, cytochemical and autoradiographical methods. The biochemical data indicate the presence of Na⁺–K⁺-ATPase, judging from the potassium dependency and, with some restrictions, the inhibition by ouabain. Cytochemically and autoradiographically, the enzyme could be localized on the cytoplasmic surfaces of the lateral plasma membranes and the basal membrane infoldings (basal labyrinth) of the folded epithelium of the branchial heart appendage. The pdocytes of the peripheral zone of the organ reacted negatively. In addition to the Na⁺–K⁺-ATPase, a magnesium-activated adenosine triphosphatase (Mg²⁺-ATPase) was demonstrated in the folded epithelium, localized mainly in the mitochondria but also at the brush border and in the apical intercellular space, whereas a bicarbonate-stimulated ATPase (HCO ₃ ^– -ATPase) was present only in the mitochondria.This study was supported by the Deutsche Forschungsgemeinschaft     

Histochemical localization of cholinesterases and monoamines in the central heart of Sepia officinalis L. (Cephalopoda)

Summary The central heart of the coleoid cephalopod, Sepia officinalis, was studied using acetylcholinesterase and fluorescence histochemistry. Using histo- and cytochemical reactions, acetylcholinesterase was localized in the axolemma and axoplasm of specific cardiac nerve fibres, as well as in the sarcolemma and within the sarcotubular system of the muscle cells. Butyrylcholinesterase exhibited a different distribution, being found only in the luminal trabecular muscle layer. Glyoxylic-acid-induced fluorescence indicated the presence of catecholamines (emission maximum 470 nm) in cardiac nerve axons. These histochemical findings support the hypothesis that noradrenaline and/or dopamine and acetylcholine act antagonistically as natural transmitters. Fluorophores indicating the presence of serotonin were not observed. The present results are discussed in the light of previous pharmacological findings.     

Histochemical localization of cholinesterases and monoamines in the central heart of Sepia officinalis L. (Cephalopoda)

The central heart of the coleoid cephalopod, Sepia officinalis, was studied using acetylcholinesterase and fluorescence histochemistry. Using histo- and cytochemical reactions, acetylcholinesterase was localized in the axolemma and axoplasm of specific cardiac nerve fibres, as well as in the sarcolemma and within the sarcotubular system of the muscle cells. Butyrylcholinesterase exhibited a different distribution, being found only in the luminal trabecular muscle layer. Glyoxylic-acid-induced fluorescence indicated the presence of catecholamines (emission maximum, 470 nm) in cardiac nerve axons. These histochemical findings support the hypothesis that noradrenaline and/or dopamine and acetylcholine act antagonistically as natural transmitters. Fluorophores indicating the presence of serotonin were not observed. The present results are discussed in the light of previous pharmacological findings.     

Serotonergic regulation of the central heart auricles of Sepia officinalis L. (Mollusca, Cephalopoda)

In pharmacological bioassays on isolated isotonically suspended auricles of Sepia officinalis, the regulatory action of the neurotransmitter serotonin (5-hydroxytryptamine, 5-HT) on these autonomous contractile compartments was demonstrated. 5-HT induced concentration-dependent positive effects on frequency and tone, whereas the concentrations/response curve for the amplitude showed a biphasic course. All applied antagonists inhibited mainly the effect of 5-HT on frequency and amplitude. The chronotropic effects of 5-HT were blocked mainly by the 5-HT(1,2) antagonist methiothepin (pA(2)=8.01), the 5-HT(1a) antagonist NAN-190 (pA(2)-) and in lesser extent by the 5-HT(1,2) antagonist mianserin (pA(2)=6.81). In the presence of each antagonist applied the 5-HT action on amplitude was transformed into a positive inotropic effect with the maximum under influence of NAN-190 and the 5-HT(2,1c)-antagonist ketanserin. The auricular tone was also influenced by the antagonists and in combination with methiothepin it turned into strong negative tonotropic effect. In addition to the pharmacological bioassays, the presence of 5-HT in nerve endings within the auricle wall was demonstrated by immunohistochemical and fluorescence microscopic findings. Altogether the findings presented here confirm that 5-HT evokes excitatory effects on the autonomous contractile auricle of S. officinalis and acts obviously over different receptors, whereby a 5-HT(1)- and a 5-HT(2)-like seem to be involved.     

The localization of enzyme activities in the pancreatic appendages of Sepia officinalis L. (Cephalopoda).

In this study, enzyme activities of the pancreatic appendages of the ductus hepatoPancreas (the so-called "pancreas") in Sepia officinalis L. have been demonstrated by light and electron micicroscopical methods: Malate dehydrogenase, monoamine oxidase, acid phosphatase, beta-glucuronidase, adenosine triphosphatase and carbonic anhydrase were shown by the former, and monoamine oxidase, catalase, glutamic oxalacetic transaminase, choline esterase (non-specific), alkaline phosphatase, acid phosphatase and carbonic anhydrase by the latter technique. The correlation between enzyme activity and distribution, and the presumed function of the two pancreatic epithelia is discussed.     

Initial stages of food ingestion by Sepia officinalis (Mollusca: Cephalopoda)

The cuttlefish ingests much skeleton from the crustaceans and fish it preys upon. The skeletal pieces are relatively large and their dimensions bear a close relationship to the length of the buccal mass and diameter of the oesophagus. The structures of the buccal mass are instrumental in the breakdown of prey and orientation of long pieces of skeleton to ensure their entry into the oesophagus. Many pieces of skeletal material present in the stomach contents still have attached muscles, showing that there is little, or no, external digestion. Skeletal material may be important for long-term maintenance of young Sepia in captivity.     

Zur Struktur und Entwicklung des Riesenfasersystems erster Ordnung von Sepia officinalis L. (Cephalopoda)

Summary The differentiation and course of the first order giant nerve fibres in the medial and posterior suboesophageal lobes of the brain of Sepia officinalis is examined in different developmental stages. In earlier embryonic stages one pair of first order giant cells differentiates on each side. Later, as a normal phenomenon, one cell of each pair degenerates.The two remaining giant fibres cross in the palliovisceral lobe. On either side of the intersection one branch of the contralateral and one branch of the ipsilateral axon are connected with the second order giant fibres. This structure, which differs from that found in Loligo, apparently mediates the functional bilaterality of the giant fibre system.

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Supported by grant NONR 2100 through the Anton and Reinhard Dohrn foundation.     

The effects of biogenic monoamines and related agonists and antagonists on the isolated,perfused branchial heart of sepia officinalis L. (cephalopoda)

1. The chronotropic and inotropic effects of biogenic monoamines were examined on the isolated and perfused branchial heart of the common cuttlefish Sepia officinalis (L.).2. Adrenaline, noradrenaline and dopamine caused concentration-dependent increases in pressure amplitude, dopamine being 100-fold less potent than adrenaline and noradrenaline. The catecholamines hardly affected frequency.3. Octopamine, histamine and GABA did not influence normal heartbeat.4. Serotonin was either ineffective or produced variable responses, whereby the efficacy of the substance was not graded to the concentrations applied. It is supposed that serotonin is not involved in branchial heart regulation.5. The actions of different adrenergic agonists and antagonists indicate the presence of a myocardial adrenoceptor which closely resembles the α₁-type.     

Nitric oxide: a vasodilatatory mediator in the cephalic aorta of Sepia officinalis (L.) (Cephalopoda)

Evidence is presented that nitric oxide (NO) may regulate blood pressure in cephalopod molluscs. In vitro tests performed on the cephalic aorta of Sepia officinalis (L.) (Cephalopoda) showed that the NO releasers (glyceroltrinitrate, sodium nitroprusside, 3-morpholinylsydnoneimine chloride and KNO(2)) induced concentration-dependent vasodilatation of vessel segments (without the tunica adventitia/periadventitia) precontracted by dopamine. These vasodilatatory actions could be totally blocked by oxadiazolo[4,3-a] quinoxalin-1-one, an inhibitor of the NO-sensitive guanylyl cyclase, and partially mimicked by the cyclic guanosine monophosphate (cGMP) analogue 8-bromo cGMP and by the phosphodiesterase inhibitor, zaprinast. The NO-precursor, L-arginine, showed vasodilatatory effects only on segments of the aorta in which the layers containing nerves (tunica adventitia/periadventitia) had been left intact, suggesting that NO synthase may be located within peripheral nerves.     

Nitric oxide: a vasodilatatory mediator in the cephalic aorta of Sepia officinalis (L.) (Cephalopoda)

Evidence is presented that nitric oxide (NO) may regulate blood pressure in cephalopod molluscs. In vitro tests performed on the cephalic aorta of Sepia officinalis (L.) (Cephalopoda) showed that the NO releasers (glyceroltrinitrate, sodium nitroprusside, 3-morpholinylsydnoneimine chloride and KNO₂) induced concentration-dependent vasodilatation of vessel segments (without the tunica adventitia/periadventitia) precontracted by dopamine. These vasodilatatory actions could be totally blocked by oxadiazolo[4,3-a] quinoxalin-1-one, an inhibitor of the NO-sensitive guanylyl cyclase, and partially mimicked by the cyclic guanosine monophosphate (cGMP) analogue 8-bromo cGMP and by the phosphodiesterase inhibitor, zaprinast. The NO-precursor, l-arginine, showed vasodilatatory effects only on segments of the aorta in which the layers containing nerves (tunica adventitia/periadventitia) had been left intact, suggesting that NO synthase may be located within peripheral nerves. Accepted: 11 August 1998     

Adenosine triphosphatase localization in the branchial heart appendage of Sepia officinalis L. (Cephalopoda).

Sodium- and potassium-dependent adenosine triphosphatase (Na+--K+-ATPase) has been demonstrated in the branchial heart appendage (pericardial gland) of Sepia officinalis L. by biochemical, cytochemical and autoradiographical methods. The biochemical data indicate the presence of Na+--K+-ATPase, judging from the potassium dependency and, with some restrictions, the inhibition by ouabain. Cytochemically and autoradiographically, the enzyme could be localized on the cytoplasmic surfaces of the lateral plasma membranes and the basal membrane infoldings (basal labyrinth) of the folded epithelium of the branchial heart appendage. The pdocytes of the peripheral zone of the organ reacted negatively. In addition to the Na+--K+-ATPase, a magnesium-activated adenosine triphosphatase (Mg2+-ATPase) was demonstrated in the folded epithelium, localized mainly in the mitochondria but also at the brush border and in the apical intercellular space, whereas a bicarbonate-stimulated ATPase (HCO-3-ATPase) was present only in the mitochondria.