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The gut of silver eels (Anguilla anguilla L.) was investigated in order to describe both the cholinergic and adrenergic intramural innervations, and the localization of possible accessory neuromediators. Histochemical reactions for the demonstration of nicotinamide adenine dinucleotide phosphate, reduced form-(NADPH-)diaphorase and acetylcholinesterase (AChEase) were performed, as well as the immunohistochemical testing of tyrosine hydroxylase, met-enkephalin, substance P, calcitonin gene-related peptide (CGRP), bombesin, vasoactive intestinal peptide (VIP), neuropeptide Y (NPY), somatostatin, cholecystokinin-octapeptide (CCK-8), serotonin, cholineacetyl transferase. The results evidenced a different pattern in comparison with other vertebrates, namely mammals, and with other fish. Both NADPH-diaphorase and AChEase activities were histochemically detected all along the gut in the myenteric plexus, the inner musculature and the propria-submucosa. Tyrosine hydroxylase immunoreactivity was observed in the intestinal tract only, both in the myenteric plexus and in the inner musculature. Several neuropeptides (metenkephalin, CGRP, bombesin, substance P, VIP, NPY, somatostatin) were, in addition, detected in the intramural innervation; some of them also in epithelial cells of the diffuse endocrine system (met-enkephalin, substance P, NPY, somatostatin). Serotonin was only present in endocrine cells. Tyrosine hydroxylase immunoreactivity was present in localizations similar to those of NADPH-diaphorase-reactivity, and in the same nerve bundles in which substance P- and CGRP-like-immunoreactivities were detectable in the intestinal tract. In addition, NADPH-diaphorase-reactive neurons showed an anatomical relationship with AChEase-reactive nerve terminals, and a similar relationship existed between the latter and substance P-like immunoreactivity.

In the last year a new congenital muscular disorder, defined as “congenital pseudo-myotonia” has been described in Chianina cattle, one of the most important Italian breed for meat quality (Fiorentina steak) and leather (used in high design easy-chair manufacture). The clinical picture is mainly characterized by an exercise-induced muscle contracture which prevents animals from performing muscular activities. On the basis of clinical symptoms Chianina cattle congenital pseudo-myotonia was related to human Brody’s disease. In 1969 Brody first described a muscular disorder characterized by an “impairment of muscle relaxation” in a human patient. Brody’s disease is known as a rare inherited disorder of skeletal muscle due to a sarco(endo)plasmic reticulum Ca2+-ATPase isoform 1 (SERCA1) deficiency, resulting from a defect of ATP2A1 gene coding for SERCA1. The SERCA1, is the protein involved in transporting calcium from cytosol into SR lumen. DNA sequencing of congenital pseudo-myotonia affected calves have revealed a missense mutation in exon 6 of the ATP2A1 gene, leading to a Arg 164 His substitution in the SERCA1 protein. Here, we provide evidence of a deficiency of SERCA1 expression and activity in sarcoplasmic reticulum membranes extracted from affected muscles, although the mRNA levels result unchanged. The loss of SERCA1 protein is not compensated by the expression of slow-twitch muscle isoform SERCA2a. We believe that Chianina cattle congenital pseudo-myotonia might be the true counterpart of human Brody's disease and that bovine specie might be used as a suitable animal model.

In mammals, glial cell line-derived neurotrophic factor (GDNF) is a growth factor of many neuronal populations in the central, peripheral and autonomous nervous system. GDNF may also function as a morphogen during kidney development and may regulate spermatogonial differentiation. GDNF has been characterised in zebrafish embryos and was demonstrated experimentally to be critical for the development of the enteric nervous system. However, in adult zebrafish, no data exist regarding GDNF expression and localisation in the brain and in different organs. Thus, the aim of the present study was to investigate the expression of GDNF in the brain of adult zebrafish (Danio rerio). Transcripts of GDNF mRNA were observed in brain extracts by a standard RT-PCR. The presence of the protein in the brain homogenates was confirmed by SDS-PAGE electrophoresis and Western blotting analysis. Immunohistochemistry and in situ hybridization experiments showed that GDNF protein and mRNA were localised in various nuclei of the telencephalon, diencephalon, mesencephalon, cerebellum and medulla oblongata of the zebrafish brain. In conclusion, this study showed that the expression of GDNF was not restricted to developmental periods but it seems that this factor might be involved in adult zebrafish brain physiology, as observed in mammals.

The authors present a muscle function disorder observed in 11 Chianina animals. The clinical picture is mainly characterized by an exercise-induced muscle contracture which prevents animals from performing muscular activities more intense than a simple walk at a slow pace. In fact, when stimulated to move faster, the muscles immediately become stiff and “freeze up” temporarily, inducing a rigid and uncoordinated gait or bunny hopping. The stiffness disappears as soon as the exercise ceases. If the exercise is prolonged, the stiffness freezes the locomotion and the animals fall on the ground like a log. The animals do not pass out or lose consciousness as in fainting. Retraction of the bulbi and prominence of the third eyelid are other common findings during these startle-elicited crises. Biochemistry usually shows slightly increased levels of creatine kinase, lactate dehydrogenase, aspartate aminotransferase and L-Lactate. Standard needle electromyography showed normal spontaneous insertion activity and voluntary electrical activity. Routine histological preparations as well as more specific histochemical and histoenzymatic stains carried out on bioptic samples of the semimembranosus muscle failed to show any remarkable and helpful findings. Genealogical evaluation of the affected animals revealed common ancestors which might suggest a genetic etiology. On the basis of the clinical findings and of the scanty results obtained by collateral exams, we have adopted the term “Congenital Pseudo-myotonia”. As regards human medicine, the disease in some way resembles the so-called “Brody’s disease”.

The contraction and relaxation of skeletal muscle is regulated by Ca release from sarcoplasmic reticulum (SR). The uptake of Ca into the lumen of SR, allowing relaxation, is mediated by a sarco-endoplasmic reticulum Ca-ATPase (SERCA). In the last years, a disorder of skeletal muscle function was described in some bovine specimens of Chianina breed. Chianina is one of the most important Italian breed for meat quality (fiorentina) and leather. Synthoms affect preferentially the musculature of the limbs and are exercise-induced stiffness and impaired skeletal muscle relaxation. This pathological condition appears us very similar to a skeletal muscle disorder described for the first time by Brody in 1969 in human patients. Brody ‘s disease is a rare inherited disease associated to a mutation in the ATP2A1 gene, encoding SERCA 1 isoform. From semimembranous pathological muscle of Chianina specimen, we prepared a crude microsomes membrane fraction (TM) in which the main markers of junctional (RyR) and non junctional SR (SERCA) are selectively enriched. TM fraction were used for determination of the ATPase activity of the SR calcium pump, assayed by a spectrophotometer method, measuring the ossidation of NADH at 340 nm, in the presence of calcium ionophore A23187. By comparison with control bovine muscle of the same species, ATPase activity of SERCA in pathological muscles resulted reduced of about 50% in two specimens and almost absent in other subjects. Immunoblotting results with antibodies against SERCA 1 isoform in the same fractions, confirmed the absence or reduction of expression of the Ca-pump in pathological animals. Our preliminary data suggests that the bovine skeletal muscle disorder might be related to human Brody’s disease and that bovine might be assumed as the animal model as porcine for malignant hyperthermia.

Electron-microscopic examinations of the sturgeon gut were performed. Oesophageal goblet cells were abundant in the stratified epithelium. The ultrastructural features of the secretory granules of the oesophageal and intestinal goblet cells were quite similar to those of other vertebrates. Lobules of multilocular adipose tissue were observed in the deep tunica propriasubmucosa of the oesophagus, in close association with vasculature and large fibre bundles of myelinated and unmyelinated axons. Similarly composed nerve fibre bundles were observed in the cardiac stomach. too. The presence of myelinated axons is an unusual feature in the vertebrate enteric nervous system. Cardiac and fundic zones of the stomach showed an epithelium with columnar ciliated and non-ciliated cells, the latter equipped with fuzzy microvilli. Cells lining the tubular gastric proper glands were markedly granulated. Intestinal superficial epithelium was columnar and contained ciliated, as well as non-ciliated and goblet cells. In the tunica propria all over the intestine, the presence and ultrastructure of granulated cells was in addition described. Intraepithelial granulated leukocytes were seen throughout the alimentary canal. Various types of endocrine cells were seen both in the stomach and in the intestine, the size of their granules was measured and their ultrastructure described and compared to that of mammalian cell types.

Using a variety of histochemical methods we have investigated the mATPase reaction of skeletal muscle fibres in the dog. Types I, IIA, IIDog (peculiar to the dog) and IIC fibres were identified. The results reveal that the interpretation of the fibre type composition depends on the methods used.

The postnatal development of skeletal muscles was studied in growing pigs from 8 to 210 d of age. Indirect immunoperoxidase staining of frozen sections of porcine semimembranosus muscle and longissimus muscle revealed a distinct population of small fibers (tertiary myotubes) that were stained specifically by an antibody (anti-NE) selective for the developmental (embryonic and neonatal) isoforms of muscle myosin. At 8 d of age the other larger fibers were already anti-NE negative and differentiated into Types I and II. A gradual decrease in the number of anti-NE positive fibers together with a gradual increase in area of the remaining positive fibers was observed throughout the pigs' growth. These results may indicate that hyperplastic growth does not cease at birth. Possible mechanisms to explain the origin of these tertiary myotubes containing developmental isoforms of myosin are suggested.

In previous studies of middle ear muscles, the classification of fibre types by histochemical methods was particularly difficult in the bovine and porcine tensor tympani muscle, suggesting the presence of immature fibres. We therefore reexamined the tensor tympani from pigs and cattle of various ages immunohistochemically, using a panel of antimyosin antibodies, including one (anti-NE) specific for neonatal and embryonic myosins. Fibres positive to anti-NE were found in tensor tympani in both species in all ages examined; only a few of these fibres reacted exclusively with this antibody; some also contained slow myosin and the majority also contained adult fast (type IIA) myosin. Furthermore, although the remaining fibres included some of the classical types I and IIA, the majority of them showed a mismatch between their histochemical and immunohistochemical profiles. The morphological appearance of the muscle, the widespread presence of neonatal myosin (often together with another myosin in the same fibre) and the persistence of this composition from birth to adulthood, could be explained by an incomplete development of the muscle fibres, resulting in a 'muscle' much better suited to the role of a ligament.