The structure and ultrastructure of the sinus venosus in the mature dogfish (Scyliorhinus canicula): the endocardium, the epicardium and the subepicardial space

The sinus venosus of fish is the most caudal chamber of the heart. It is often reduced in teleosts but well developed in elasmobranchs. The sinus venosus of the dogfish (Scyliorhinus canicula) is vital, since it harbours key elements such as a little known neuroendocrine system and the nodal tissue. However, the study of its structure is still incomplete. We examined the endocardium, epicardium and subepicardium of the sinus venosus in mature dogfishes. The wall is 100-250 microm thick and comprises three main layers. Large bundles of myocardial cells occupy the middle layer. The endothelial ensheathing is composed of thin endocardial cells with prominent nuclei towards the lumen, whose cytoplasm contains numerous dense bodies and moderately dense bodies, 150-800 nm in diameter and large vacuoles. The possible functions of these organelles are discussed. The outermost layer is made of a robust sheet of cuboidal epicardial cells separated from the subepicardium by a conspicuous basal lamina. Numerous microvilli towards the pericardial cavity and elliptical vesicles are located in the apex of epicardial cells. A thick layer richly endowed with dense bundles of collagen fibres forms the subepicardial space. This structure should be contrasted with the venous return mechanism of elasmobranchs.     

Mechanism of uptake and retrograde axonal transport of noradrenaline in sympathetic neurons in culture: reserpine-resistant large dense-core vesicles as transport vehicles

The uptake and retrograde transport of noradrenaline (NA) within the axons of sympathetic neurons was investigated in an in vitro system. Dissociated neurons from the sympathetic ganglia of newborn rats were cultured for 3-6 wk in the absence of non-neuronal cells in a culture dish divided into three chambers. These allowed separate access to the axonal networks and to their cell bodies of origin. [3H]NA (0.5 X 10(-6) M), added to the axon chambers, was taken up by the desmethylimipramine- and cocaine-sensitive neuronal amine uptake mechanisms, and a substantial part was rapidly transported retrogradely along the axons to the nerve cell bodies. This transport was blocked by vinblastine or colchicine. In contrast with the storage of [3H]NA in the axonal varicosities, which was totally prevented by reserpine (a drug that selectively inactivates the uptake of NA into adrenergic storage vesicles), the retrograde transport of [3H]NA was only slightly diminished by reserpine pretreatment. Electron microscopic localization of the NA analogue 5-hydroxydopamine (5-OHDA) indicated that mainly large dense-core vesicles (700-1,200-A diam) are the transport compartment involved. Whereas the majority of small and large vesicles lost their amine dense-core and were resistant to this drug. It, therefore, seems that these vesicles maintained the amine uptake and storage mechanisms characteristic for adrenergic vesicles, but have lost the sensitivity of their amine carrier for reserpine. The retrograde transport of NA and 5-OHDA probably reflects the return of used synaptic vesicle membrane to the cell body in a form that is distinct from the membranous cisternae and prelysosomal structures involved in the retrograde axonal transport of extracellular tracers.     

Endocardial pores in the sinus venosus and atrium of the dogfish

A scanning electron microscopy study showed the presence of large pores in the endocardium of the sinus venosus of the dogfish Scyliorhinus canicula. The pores were always found on large bundles which protruded into the cardiac lumen. The bundles were mainly constituted of granule-containing nerve fibres. The average diameter of the pores was 3.2 μm (range=1.5-5.0μm), and their density, was about 822 pores mm⁻². Endocardial pores were absent in other areas of the sinus venosus, but they were observed on the little bundles of granulated nerve fibres which were scattered throughout the atrium. The existence of large endocardial pores associated with bundles of granulated nerve fibres supports the hypothesis for the neuroendocrine nature of the elasmobranch sinus venosus wall.     

Studies of cardiac ganglia in pre- and postnatal rabbits

Summary This investigation was undertaken to describe the ultrastructure of cardiac ganglia in rabbits from day 18 of gestation to day 35 postpartum. Special attention was directed to the types of synaptic contacts made with the principal neurons and with the small granule-containing cells. The cardiac ganglia in all animals consisted mainly of parasympathetic postganglionic neurons, supporting cells, and small granule-containing (small intensely fluorescent) cells. The neurons received afferent synaptic terminals of two types. One type contained mainly small clear vesicles typical of most cholinergic terminals. The second type contained mainly small dense-core vesicles (these were most prominent after treatment of the animal with 5-hydroxydopamine), and were considered to be adrenergic terminals. These adrenergic terminals are probably part of an inhibitory system in the ganglia. The small granule-containing cells received typical afferent synaptic terminals of the cholinergic type, and also formed specialized contacts with certain axonal terminals. These latter specializations are considered to be reciprocal synapses which probably have a role in modulating ganglionic transmission.Supported by the Kentucky Heart Association and the Heart Association of Louisville and Jefferson County     

Tachykinin-like immunoreactivity in the sinus venosus of the dogfish (Scyliorhinus canicula)

The sinus venosus of the elasmobranch heart is characterized by the presence of large bundles of unmyelinated nerve fibres that bulge into the cardiac lumen, below the endocardium. In the dogfish (Scyliorhinus canicula), these fibres contain numerous dense-core membrane-bounded granules of about 200 nm in diameter. Most intramural ganglion cells of the sinus venosus also show densely packed granules similar to those found in the subendocardial fibres. We have observed strong substance-P-like immunoreactivity in the large fibre bundles and in the perikarya of the ganglion cells. Preabsorption of the antisera with fragment 7–11 of substance P has shown that the antisera recognize the tachykinin canonic sequence. Our findings suggest that an undetermined tachykinin is secreted in the elasmobranch heart, and that it is probably released into the blood stream in the context of a little-known neuroendocrine system.     

On the ultrastructure of the sinus venosus in Chimara monstrosa L. (Elasmobranchii: Holocephali)

Summary The wall of the sinus venosus in an elasmobranchian species, Chimaera monstrosa L. is described.Endocardial cells contain numerous large vacuoles, as well as a number of membrane-bounded, moderately electron dense bodies (MDB). Myocardial cells lie closely packed into bundles surrounded by a basal lamina of about 20 nm thickness, and by large amounts of collagen fibres. These cells are connected by desmosomes of 1–2 µm length and with an intermembranous gap of 10–20 nm. Myocardial cells poor in myofibrils are intermingled with cells containing a well developed contractile material. Atrial specific granules are scarce. Vesiculated nerve processes occur at a distance of about 20 nm from the myocardial sarcolemma. Myocardial cells of the sino-atrial junction appear ultrastructurally similar to those located elsewhere in the sinus venosus. Epicardial cells contain large vacuoles, and have fibrecoated protrusions extending into the pericardial space.The possibility of pacemaker activity in the elasmobranchian sinus venosus is discussed.     

Ultrastructural study of the myocardium of the sinus venosus and sinoatrial valve in the dogfish (Scyliorhinus canicula)

The myocardium of the sinus venosus of the dogfish ( Scyliorhinus canicula ) is located between a thick subepicardial collagen-rich layer and a subendocardial network of nerve fibres and ganglion cells. The sinoatrial valve consists of two transversal folds of the cardiac wall which are separated by connective tissue, except in their free margins.
The myocardium of the sinus venosus and the sinusal face of the sinoatrial valve is arranged in bundles which are surrounded by a 40 nm-thick basal lamina. The myocardial cells measure about 7-9 μm in diameter at the nuclear level. Nerve terminals are frequent in the centre of the bundles. Most of the sinusal myocardiocytes have a scarce amount of myofibrils which are randomly orientated. The sarcoplasmic reticulum is relatively well developed and consists of peripheral couplings, subsarcolemmal vesicles, circular and longitudinal tubules. The scarce intercalated discs show only fasciae adhaerentes . Gap junctions, desmosomes or specific granules are not observed in the sinusal myocardiocytes of the dogfish. In contrast, the atrial myocardiocytes are smaller, about 5-6 μm in diameter at the nuclear level. The cytoplasm is denser and the myofibrils are abundant and orientated in parallel directions. Specific granules are present. although scarce. Subsarcolemmal vesicles are less frequent, while the atrial intercalated discs are larger and more abundant than those of the sinus venosus. Neural elements are scarce in the atrium.
The differences observed between sinus venosus and atrium might be related to the morphological criteria to distinguish between the nodal tissue and the working myocardiocytes of higher vertebrates. On the other hand, we think that the connective tissue placed between sinus venom and atrium means that the contraction impulse generated in the sinus venosus must reach the atrium through the free margin of the valve. It might play a role in the sinoatrial valve function.     

The innervation and fine structure of paraneuronic cells in an amphibian pulmonary artery

Summary The pulmonary artery of Bufo marinus contains large numbers of bipolar cells situated in the tunica adventitia and in the outer layers of the media. These cells show a bright green-yellow fluorescence (emission spectra 485 nm) after formaldehyde pre-treatment suggesting that they contain a primary monoamine. The most characteristic fine-structural feature of these cells is the presence of numerous dense-cored vesicles (80—300 nm diameter) in their cytoplasm. The cells are in close contact (20 nm gap) with both agranular and granular nerve fibres. Both EM-cytochemical and formaldehyde-induced fluorescence tests indicate that the granule-containing nerve fibres are adrenergic. The agranular nerve fibres form discrete synaptic contacts with pre-and post-synaptic membrane thickenings on the cells. This was never observed with respect to the adrenergic fibres. Each process of the cells is about 45 m long. The processes do not bear any special relationship to either vessels of the arterial vasa vasorum or medial smooth muscle cells. Their location in the wall of the artery suggests that they are functionally significant with respect to activity of the arterial media.     

Ultrastructure of sympathetic ganglion cells and granule-containing cells in the paracervical (Frankenhäuser) ganglion of the newborn rat

Summary A study was made of the ultrastructure of the paracervical (Frankenhäuser) ganglion of the newborn rat, using immersion fixation by glutaraldehyde (2.5%) followed by OsO₄ (1%), or KMnO₄ (3%) fixation. The cells containing dense—core vesicles were divided into three groups: (1) primitive sympathetic cells, (2) cells containing some dense-core vesicles 700–1100 Å in size and structurally resembling sympathetic neurons, called principal neurons, and (3) cells containing many dense-core vesicles with a larger, darker dense core, 800–2000 Å in diameter, called granule-containing cells. Using glutaraldehyde-osmium fixation, the principal neurons were further divided into dark and light cells on the basis of electron opacity of the cytoplasmic matrix. The granule-containing cells were believed to correspond to the small, intensely fluorescent cells (SIF-cells) previously described using the formaldehyde-induced fluorescence technique. On the basis of the amount of granules, the granulecontaining cells were classified as mature or maturing SIF-cells and as more primitive SIF-cells, and developing sympathicoblasts. The development of synapses in autonomic ganglia was discussed.Grant: The Finnish Medical Foundation.     

Heart inflow tract of the African lungfish Protopterus dolloi

We report a morphologic study of the heart inflow tract of the African lungfish Protopterus dolloi. Attention was paid to the atrium, the sinus venosus, the pulmonary vein, and the atrioventricular (AV) plug, and to the relationships between all these structures. The atrium is divided caudally into two lobes, has a common part above the sinus venosus, and appears attached to the dorsal wall of the ventricle and outflow tract through connective tissue covered by the visceral pericardium. The pulmonary vein enters the sinus venosus and runs longitudinally toward the AV plug. Then it fuses with the pulmonalis fold and disappears as an anatomic entity. However, the oxygenated blood is directly conveyed into the left atrium by the formation of a pulmonary channel. This channel is formed cranially by the pulmonalis fold, ventrally by the AV plug, and caudally and dorsally by the atrial wall. The pulmonalis fold appears as a wide membranous fold which arises from the left side of the AV plug and extends dorsally to form the roof of the pulmonary channel. The pulmonalis fold also forms the right side of the pulmonary channel and sequesters the upper left corner of the sinus venosus from the main circulatory return. The AV plug is a large structure, firmly attached to the ventricular septum, which contains a hyaline cartilaginous core surrounded by connective tissue. The atrium is partially divided into two chambers by the presence of numerous pectinate muscles extended between the dorsal wall of the atrium and the roof of the pulmonary channel. Thus, partial atrial division is both internal and external, precluding the more complete division seen in amphibians. The present report, our own unpublished observations on other Protopterus, and a survey of the literature indicate that not only the Protopterus, but also other lungfish share many morphologic traits.     

A light and electron microscopic study on the embryonic development of the rat carotid body.

The embryonic development of the rat carotid body was studied with electron microscopy. In the 11 mm embryo a cell aggregation consisting of undifferentiated cells and unmyelinated nerve fibers appears on the anterior wall of the third branchial artery. Granule-containing cells appear in the 12 mm embryo and continue to increase in number as the cellular aggregation increases in size and becomes separated from the wall of the third branchial artery. Synapse formation and the appearance of fenestrated capillaries occur almost simultaneously at the 17 mm stage. There are two types of synapses, one with membrane densification and vesicles clustered inside the nerve endings, the other with dense material and vesicles inside the granule-containing cells. At the 20 mm stage the undifferentiated cells send enveloping cytoplasmic processes toward adjacent granule-containing cells and the carotid body anlage displays rudimentary lobules.     

The ultrastructure of paraganglia associated with the inferior mesenteric ganglia in the guinea-pig

Summary The paraganglia of the inferior mesenteric ganglia in the guinea-pig are composed of small chromaffin cells containing an abundance of granule-containing vesicles. The chromaffin cells are almost completely surrounded by satellite cells. In areas in which satellite cell processes do not intervene, the membranes of adjacent chromaffin cells are closely apposed and often form specialized attachment zones. The paraganglia contain a dense capillary network, the endothelial cells of which are often extremely attenuated and show areas of fenestration. The processes of chromaffin cells approach close to the capillary walls and are often bare of satellite cells covering on the side facing the capillary. Evidence has been obtained for the exocytotic release of the contents of chromaffin cell vesicles into pericapillary spaces. Synapses of cholinergic and noradrenergic axons are seen on the chromaffin cells. The cholinergic axons degenerate when the praganglia are decentralized, but the noradrenergic axons, which appear to arise from the local inferior mesenteric ganglia, remain intact. The results suggest that the paraganglia have an endocrine function.     

Differentiation of embryonic chick sympathetic neurons in vivo: ultrastructure,and quantitative determinations of catecholamines and somatostatin

Summary The ultrastructural and transmitter development of lumbar sympathetic ganglia was studied in embryonic day-6 through-18 chick embryos. At embryonic day 6, ganglia are populated by two morphologically distinct types of neuronal cells and Schwann cell precursors. The neuronal populations basically comprise a granule-containing cell and a developing principal neuron. Granule-containing cells have, an irregularly shaped or oval nucleus with small clumps of chromatin attached to the inner nuclear membrane and numerous large (up to 300 nm) membrane-limited granules. Developing principal neurons display a more rounded vesicular nucleus with evenly distributed chromatin, prominent nucleoli, more developed areas of Golgi complexes, and rough endoplasmic reticulum and large dense-core vesicles up to 120 nm in diameter. There are granule-containing cells with fewer and smaller granules which still display the nucleus typical for granule-containing cells. These granule-containing cells may develop toward developing principal neurons or the resting state of granule-containing cells found in older ganglia. Both granule-containing cells and developing principal neurons proliferate and can undergo degeneration. At embryonic day 9 there are far more developing principal neurons than granule-containing cells. Most granule-containing cells have very few granules. Mitotic figures and signs of cell degeneration are still apparent. Synapse-like terminals are found on both developing principal neurons and granule-containing cells. Ganglionic development from embryonic day 11 through 18 comprises extensive maturation of developing principal neurons and a numerical decline of granule-containing cells. Some granule-containing cells with very few and small granules still persist at embryonic day 18. The mean catecholamine content per neuron increases from 0.044 femtomol at embryonic day 7 to 0.22 femtomol at embryonic day 15. Concomitantly, there is a more than 6-fold increase in tyrosine hydroxylase activity. Adrenaline has a 14% share in total catecholamines at embryonic day 15. Somatostatin levels are relatively high at embryonic day 7 (1.82 attomol per neuron) and are 10-fold reduced by embryonic day 15. Our results suggest the presence of two morphologically distinct sympathetic neuronal precursors at embryonic day 6: one with a binary choice to become a principal neuron or to die, the other one, a granule-containing cell, which alternatively may develop into a principal neuron, acquire a resting state or die.     

The innervation of frog's taste organ “A histochemical study”

Morphological investigations on the fungiform papilla of the frog (Rana pipiens) have shown that this taste organ contains two distinct populations of cells: associate and sensory. Messages received by the sensory cells are believed to be transmitted through the mediation of an adrenergic transmitter. This chemical was shown by fluorescence microscopy and electron histochemistry to be stored in synaptic granular vesicles which accumulate at the membrane of the cytoplasmic processes of the sensory cells in typical chemical synaptic complexes. The sensory cell cytoplasmic processes form the presynaptic component of these complexes whose post synaptic components are the nerve fibres supplying the taste buds. These sensory nerve fibres contain agranular vesticles and are probably cholinergic, since they show positive cholinesterase activity at the light and electron microscopical levels.     

Different types of small granule-containing cells and neurons in the guinea-pig adrenal medulla

Summary An electron microscopic, histoand biochemical study was carried out on the adrenal medulla of newborn and adult guinea-pigs giving special emphasis to small granule-containing (SGC) cells. Adrenaline (A) was the predominating catecholamine (CA) both in newborn (70–90 % of total CA) and adult (85–90%) guinea-pig adrenals. In analogy to the biochemical findings electron microscopy revealed a high predominance of A cells, which contained large granular vesicles with an average diameter of 180 nm. Most noradrenaline (NA) storing cells showed granular vesicles of a considerably smaller average diameter (80 nm) and had a higher nuclear-cytoplasmic ratio. These cells were termed SGC-NA cells. NA cells with large granular vesicles (average diameter 170 nm) were extremely rare. Another type of SGC cells contained granular vesicles with cores of low to medium electron-density (SGC-NA-negative cells). Biochemical determinations made it unlikely that these cells contained predominantly dopamine (DA). SGC cells were scarcely innervated by cholinergic nerves. They formed processes, which were found both in the adrenal cortex and medulla contacting blood vessels including sinusoid capillaries, steroid producing cells of the reticularis and fasciculata zone and processes, which were interpreted to belong to medullary nerve cells.Two types of neurons were present in the guinea-pig adrenal medulla, one resembling the principal neurons in sympathetic ganglia, the other, which, according to its morphology, occupied an intermediate position between principal neurons and SGC cells.In adrenomedullary grafts under the kidney capsule, which were studied three weeks after transplantation, ordinary A cells resembled SGC-NA negative cells with respect to their ultramorphology. Processes of transplanted principal neurons showed uptake of 5-hydroxydopamine and, hence, were considered to be adrenergic. Despite the lack of extrinsic nerves to the transplants, few principal neurons received cholinergic synapses, the origin of which is uncertain to date.Supported by a grant from Deutsche Forschungsgemeinschaft (Un 34/4)Dedicated to Professor H. Leonhardt in honor of his 60th birthday.     

Dense-core vesicles and non-synaptic exocytosis in the central body of the crayfish brain

Summary The central body in the median protocerebrum of the brain of the crayfish Cherax destructor is a distinctive area of dense neuropile, the nerve fibres of which contain three main types of vesicles: electronlucent vesicles (diameter 35 nm), dense-core vesicles (diameter 64 nm), and large structured dense-core vesicles (diameter 98 nm, maximum 170 nm). Different vesicle types were found together in the same neurons. Electronlucent vesicles were seen at presynaptic sites and rarely observed in the state of exocytosis. Exocytosis of densecore and structured dense-core vesicles was a regular feature on non-synaptic release sites either close to, or at some distance from pre- and subsynaptic sites. Non-synaptic exocytotic sites are more often observed than chemical synapses. Different forms of exocytosis seen at non-synaptic sites included the release of single densecore vesicles, packets of dense-core vesicles, and rows of dense-core vesicles lined up along cell membranes and around fibre invaginations. Swelling and the enhanced electron density of extracellular non-synaptic spaces may mark the positions of prior exocytotic events. In vitro treatment of the brain with tannic acid buffer solution followed by conventional double fixation resulted in the augmentation of non-synaptic exocytosis. Electron microscopy of proctolin- and serotonin-immunoreactive nerve fibres shows them to contain dense-core and electron-lucent vesicles and to be surrounded by many unlabelled profiles similarly laden with dense-core vesicles and electron-lucent vesicles, indicating the presence of other, not yet identified, neuroactive compounds.     

Structure and innervation of the pineal gland of the rabbit,Oryctolagus cuniculus (L.)

In the rabbit pineal gland two types of postganglionic nerve endings were found which are characterized by the presence of small dense-core vesicles or small clear vesicles. Pharmacological and cytochemical experiments showed then to be noradrenergic and cholinergic, respectively. Both types were often present in the same nerve bundle, occasionally in close opposition. Intrapineal neurons were only rarely observed. They showed cholinergic synapses on their perikaryon and dendrites as well as noradrenergic axo-dendritic close contacts. Bilateral extirpation of the superior cervical ganglia revealed the postganglionic sympathetic origin of the pineal noradrenergic nerve fibres. Moreover, it appeared that these ganglia are hardly, if at all, involved in the pathway of pineal cholinergic innervation. The results obtained from lesions of both facial nerves, taken together with the results reported in the literature, led to the conclusion that the postganglionic cholinergic nerve fibers in the pineal are of parasympathetic origin. A model for the sympathetic and parasympathetic pineal innervation is proposed.     

Immunocytochemical and electron-microscopic study of the elasmobranch nucleus sacci vasculosi

Summary The elasmobranch nucleus sacci vasculosi was studied by means of electron microscopy (in the dogfish) and immunocytochemistry (in the dogfish and the skate) by using antibodies against tyrosine hydroxylase, alpha-melanocyte-stimulating hormone, somatostatin, serotonin, and substance P. Ultrastructural study of the dogfish nucleus sacci vasculosi shows the presence of medium-sized cells that possess numerous mitochondria but that have no dense-core vesicles in the cytoplasm or in cell processes. Fibres of the conspicuous tractus sacci vasculosi have a beaded appearance and form conventional synapses with dendrites and cell perikarya of the nucleus sacci vasculosi. The perikarya of this hypothalamic nucleus were not immunoreactive to any of the antibodies tested, and fibres immunopositive to tyrosine hydroxylase, alpha-melanocyte-stimulating hormone, somatostatin, serotonin, and substance P were scarce within this nucleus, in both the dogfish and the skate. Dorsal to the nucleus sacci vasculosi, there are numerous positive neuronal processes in addition to many small neurons that show immunoreactivity to alpha-melanocyte-stimulating hormone, somatostatin and tyrosine hydroxylase. Two types of neuron occur in this dorsal region, displaying dense-core vesicles of either 100–160 nm or 60–100 nm diameter in their cytoplasm; they were identified as peptide-containing and monoamine-containing neurons, respectively. The neuropil of this region has a significantly different ultrastructure from that of the nucleus sacci vasculosi, with many processes containing dense-core vesicles. This group of neurons, located dorsal to the nucleus sacci vasculosi and showing (a) immunoreactivity to neuropeptides or to monoamine-synthesizing enzyme, and (b) cytoplasm with dense-core vesicles, was considered not to be a part of the nucleus sacci vasculosi but rather part of the nucleus tuberculi posterioris. These results support the non-peptidergic and non-aminergic character of the nucleus sacci vasculosi.     

Subcellular localization of chromogranins, calcium channels, amine carriers, and proteins of the exocytotic machinery in bovine splenic nerve

Subcellular fractionation of bovine splenic nerves, which consist mainly of sympathetic nerve fibers, has been useful for characterizing cellular organelles en route to the terminal. In the present study we have characterized the subcellular distribution of both secretory and membrane proteins. A newly discovered chromogranin-like protein, NESP55, was found in large dense-core vesicles. The endogenous processing of NESP55 was comparable to that of chromogranins but more limited than that of secretogranin II and chromogranin B. For membrane proteins three major types of distribution were found. The amine carrier VMAT2 was confined to large dense-core vesicles. VAMP or synaptobrevin was present both in large dense-core vesicles and in lighter vesicles, whereas SNAP-25, syntaxin, and two types (N and L) of Ca2+ channels were found in a special population of lighter vesicles but were not present in large dense-core vesicles or at the most in very low concentrations. The plasma membrane norepinephrine transporter was apparently present in a separate type of vesicle, but this requires further study. These results further characterize vesicles en route to the terminal and establish for the first time that peptides involved in exocytosis (syntaxin, SNAP-25, and N- and L-type Ca2+ channels) are apparently transported to the terminal in a special type of vesicle. The exclusive presence of the amine carrier in large dense-core vesicles indicates that the formation of small dense-core vesicles in the terminals requires a reuse of membrane components of large dense-core vesicles.     

Ultrastructure and cytochemistry of secretory cells in the skin of the leech,Dina lineata

The ultrastructure and histochemical features of the two types of secretory cells in leech skin are described. Pear-shaped cells secrete mucus containing carboxylated mucosubstances, while tubular cells produce a mucus containing a mixture of neutral, carboxylated, and sulfated mucosubstances. Pear-shaped secretory cells have two types of neuroglandular junctions, one containing dense-core serotonergic vesicles and the other small clear vesicles. Tubular secretory cells have large terminals, with many clear vesicles thought to be cholinergic. © 1993 Wiley-Liss, Inc.