Catecholaminergic innervation of muscles in the hindgut of crustaceans

Summary The crustacean species Pacifastacus leniusculus and Gammarus pulex were investigated by electron microscopy in a search for possible neuromuscular junctions in the hindgut, which has a rich supply of catecholaminergic fibres. True neuromuscular synapses were found in both species between nerve terminals containing dense-core vesicles (80–110 nm in diam.) and muscle fibres. We suggest that the dense-core vesicle terminals contain a catecholamine, and this is supported by ultrahistochemical tests for monoamines. Two types of junctions are found: one in which the nerve terminal is embedded in the muscle cell (both species) and one in which protrusions from the muscle cell meet nerve terminals (Pacifastacus). Gammarus pulex, which has only circular muscles in the hindgut, has only catecholaminergic innervation, whereas Pacifastacus leniusculus has circular and longitudinal muscles both with at least two types of innervation.The investigation was supported by grants from the Swedish Natural Science Research Council (B 2760-009), the Hungarian Academy of Sciences, the Royal Swedish Academy of Sciences, and the Magnus Bergvall Foundation. We are also indebted to Mrs. Lena Sandell for her skilful technical assistance     

Ultrastructural,fluorescence microscopic and microfluorimetric study of the innervation of the axial complex in the sea urchin,Sphaerechinus granularis (Lam.)

Summary In juxtaposition with the contractile epithelia of the axial complex of the sea urchin, Sphaerechinus granularis, several types of nerve fibers with different vesicle populations were determined. Nerve terminals, filled with clear vesicles and dense core vesicles, form synaptoid neuromuscular junctions. Close to the somatocoelic epithelia of the axial and terminal sinus septa, numerous axon profiles form a nerve plexus. Among the epithelial cells covering the plexus, two types of nerve cells can be distinguished which presumably produce neurosecretory and aminergic granules, respectively. Monoamine fluorescence (formaldehyde-induced fluorescence, Falck-Hillarp technique) was analyzed microspectrofluorimetrically. The emission spectrum of the fluorophores occurring in the present material shows a maximum at 475 nm and is characteristic of catecholamines; the excitation maximum at 380 nm after formaldehyde treatment is typical of catecholamines at low pH only. Since the peak ratio (370:320 nm) does not change after HCl-vapor treatment, the fluorophores are likely to be indicative of dopamine.     

Fine structural studies of the nervous system and the apical organ in the planula larva of the sea anemone Anthopleura elegantissima

The nervous system of the planula larva of Anthopleura elegantissima consists of an apical organ, one type of endodermal receptor cell, two types of ectodermal receptor cells, central neurons and nerve plexus. Both interneural and neuromuscular synapses are found in the nerve plexus. The apical organ is a collection of about 100 long, columnar cells each bearing a long cilium and a collar of about 10 microvilli. The cilia of the apical organ are twisted together to form an apical tuft. The ciliary rootlets of the apical organ cells are extremely long, reaching to the basal processes of the cells adjacent to the mesoglea. All three types of sensory cells are tall and slender in profile and are identified by the presence of one or more of the following features: microtubules, small vesicles, membrane-bound granules and synapses. The interneurons are bipolar cells with somas restricted to the aboral end, adjacent to the apical organ. All synapses observed are polarized or asymmetrical. A diagram including all the elements of the nervous system is presented and the possible functions of the nervous system are discussed in relation to larval behavior.     

Ultrastructure of nerve plexus in flatworms

Summary Synaptic components from the peripheral nervous system of the polyclad flatworm, Notoplana acticola, are described from electron microscopic observations. Quasineuropile, defined as clusters of neurites containing synaptic vesicles, occurs as scattered islands along the peripheral nerve cords of the plexus. Some neurite clusters only contain one type of synaptic vesicle but others are mixed. The most usual synaptic configuration consists of a single presynaptic element and a pair of postsynaptic neurites sharing a common synaptic cleft. These synapses are polarized and contain clear, 420 Å vesicles. GABA-type synapses are also found. At least two kinds of solid-core vesicles also occur.     

Reduced Expression of the Vesicular Acetylcholine Transporter and Neurotransmitter Content Affects Synaptic Vesicle Distribution and Shape in Mouse Neuromuscular Junction

In vertebrates, nerve muscle communication is mediated by the release of the neurotransmitter acetylcholine packed inside synaptic vesicles by a specific vesicular acetylcholine transporter (VAChT). Here we used a mouse model (VAChT KD^HOM) with 70% reduction in the expression of VAChT to investigate the morphological and functional consequences of a decreased acetylcholine uptake and release in neuromuscular synapses. Upon hypertonic stimulation, VAChT KD^HOM mice presented a reduction in the amplitude and frequency of miniature endplate potentials, FM 1–43 staining intensity, total number of synaptic vesicles and altered distribution of vesicles within the synaptic terminal. In contrast, under electrical stimulation or no stimulation, VAChT KD^HOM neuromuscular junctions did not differ from WT on total number of vesicles but showed altered distribution. Additionally, motor nerve terminals in VAChT KD^HOM exhibited small and flattened synaptic vesicles similar to that observed in WT mice treated with vesamicol that blocks acetylcholine uptake. Based on these results, we propose that decreased VAChT levels affect synaptic vesicle biogenesis and distribution whereas a lower ACh content affects vesicles shape.     

Nerve terminals and synapses in the cardiac ganglion of the adult lobster Homarus americanus

The cardiac ganglion in the lobster Homarus americanus was examined with a transmission electron microscope. Nerve terminals often existed in large aggregations surrounded by glial and connective tissue elements. Axo-axonic and axo-dendritic synapses were present. Six ultrastructurally different types of nerve terminal, each containing an abundance of vesicles, were distinguished: three formed discrete chemical synapses as indicated by typical release site morphology; three did not. The latter appear to be neurosecretory axon terminals of extrinsic neurons. More than one morphologically distinct type of synaptic vesicle occurred commonly in a given terminal, suggesting the presence of coexisting neurotransmitters and/or neuroregulatory factors. Symmetrical chemical synapses and electrotonic junctions between axons were present.     

Neural Organization of the Median Ocellus of the Dragonfly : II. Synaptic structure

Two types of presumed synaptic contacts have been recognized by electron microscopy in the synaptic plexus of the median ocellus of the dragonfly. The first type is characterized by an electron-opaque, button-like organelle in the presynaptic cytoplasm, surrounded by a cluster of synaptic vesicles. Two postsynaptic elements are associated with these junctions, which we have termed button synapses. The second synaptic type is characterized by a dense cluster of synaptic vesicles adjacent to the presumed presynaptic membrane. One postsynaptic element is observed at these junctions. The overwhelming majority of synapses seen in the plexus are button synapses. They are found most commonly in the receptor cell axons where they synaptically contact ocellar nerve dendrites and adjacent receptor cell axons. Button synapses are also seen in the ocellar nerve dendrites where they appear to make synapses back onto receptor axon terminals as well as onto adjacent ocellar nerve dendrites. Reciprocal and serial synaptic arrangements between receptor cell axon terminals, and between receptor cell axon terminals and ocellar nerve dendrites are occasionally seen. It is suggested that the lateral and feedback synapses in the median ocellus of the dragonfly play a role in enhancing transients in the postsynaptic responses.     

The Ultrastructure of the Nervous System of Gyratrix hermaphroditus (Turbellaria, Rhabdocoela)

The peripheral nervous system and the synapses of G. hermaphroditus are studied with the electron microscope. There is a submuscular as well as a subepithelial plexus. The subepithelial plexus is found among the muscles and between the muscles and the basement membrane. It consists of fibres containing large lucent and lysosome-like vesicles and fibres with only small lucent (synaptic) vesicles. In the deeper lying submuscular plexus also dense and dense-cored vesicles occur in the fibres. Cell bodies are not observed in the plexuses. The separate nerve supplies of the pharynx and the gonads contain nerve cells of the neurosecretory type. Fibres of the same kinds as in the brain are also seen here. The synapses in the neuropile are of two kinds. 1. Symmetrical synapses with an additional presynaptic network are most common. 2. Synapses without thickenings of membranes are observed between lateral membranes of neurites. In the peripheral nervous system are two other kinds of synapses also observed. 1. Asymmertical synapses with a denser and wider postsynaptic thickening and 2. neuromuscular junctions. Neurites containing accumulations of small vesicles against the basement membrane are also described. The organization of the peripheral nervous system is described and discussed in relation to the systematic position of G. hermaphroditus.     

Excitatory Transmission in Insect Neuromuscular Systems

Many, but not all, visceral muscles in insects are innervatedby neurosecretory axons. The neurosecretory junctions with theheart muscle of the American cockroach, Periplaneta americana,show ultrastructural and electrophysiological evidence of chemicallytransmitting synapses, and cytochemical evidence for the presenceof monoamines. Electron microscopy of nerve terminals showsthat synaptic vesicles may be formed directly from electron-dense"neurosecretory" granules Neurotomy of motor axons to skeletal muscles in insects leadsto aggregation and clumping of synaptic vesicles after 48 hours.Treatment of in vitro nerve-muscle preparations with variousrespiratory poisons caused aggregation similar to that developedin neurotomized animals. This suggested that vesicle aggregationin both cases may have resulted from a decrease in availableadenosine triphosphate in the nerve terminal with subsequentalteration in the normal charge density which supports a repulsiveforce between the vesicles.     

Aberrant morphology and residual transmitter release at the Munc13-deficient mouse neuromuscular synapse

In cultured hippocampal neurons, synaptogenesis is largely independent of synaptic transmission, while several accounts in the literature indicate that synaptogenesis at cholinergic neuromuscular junctions in mammals appears to partially depend on synaptic activity. To systematically examine the role of synaptic activity in synaptogenesis at the neuromuscular junction, we investigated neuromuscular synaptogenesis and neurotransmitter release of mice lacking all synaptic vesicle priming proteins of the Munc13 family. Munc13-deficient mice are completely paralyzed at birth and die immediately, but form specialized neuromuscular endplates that display typical synaptic features. However, the distribution, number, size, and shape of these synapses, as well as the number of motor neurons they originate from and the maturation state of muscle cells, are profoundly altered. Surprisingly, Munc13-deficient synapses exhibit significantly increased spontaneous quantal acetylcholine release, although fewer fusion-competent synaptic vesicles are present and nerve stimulation-evoked secretion is hardly elicitable and strongly reduced in magnitude. We conclude that the residual transmitter release in Munc13-deficient mice is not sufficient to sustain normal synaptogenesis at the neuromuscular junction, essentially causing morphological aberrations that are also seen upon total blockade of neuromuscular transmission in other genetic models. Our data confirm the importance of Munc13 proteins in synaptic vesicle priming at the neuromuscular junction but indicate also that priming at this synapse may differ from priming at glutamatergic and gamma-aminobutyric acid-ergic synapses and is partly Munc13 independent. Thus, non-Munc13 priming proteins exist at this synapse or vesicle priming occurs in part spontaneously: i.e., without dedicated priming proteins in the release machinery.