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The spatial expression pattern of Brain-Derived Neurotrophic Factor (BDNF) mRNA in the brain of the European eel (Anguilla anguilla) was determined using non-radioactive in situ hybridization, and mapped and compared to that of other vertebrates. Riboprobes were prepared based on a partial cDNA coding sequence for A. anguilla BDNF that was amplified using degenerate primers, cloned and sequenced. As in other animal groups, in the eel, BDNF mRNA expression was seen in the telencephalon, hypothalamus, tectum, many primary and secondary sensory centers, and cranial motor nuclei. However, in contrast to mammals, BDNF mRNA expression was observed in some brain stem nuclei, such as the reticular formation, that contain cell bodies of neurons that project down the spinal cord. We suggest that these differences might relate to the continual growth of teleost fish.

We describe here the meningeal sheath that encloses the spinal cord, and the sheath that develops when the cord regenerates after a total transection. This description is derived from electron and light microscopy. The sheath of the uninjured cord was found to be a single structure of two parts: an outer, thin melanocyte layer and an inner, thicker layer of 2 to 10 rows of fibroblasts, closely associated with collagen and elastic fibers. Soon after cord transection, the injured axons re-grow and, together with the reforming central canal, create a bridge that links the transected cord within 8 days of injury. This bridge is covered at first by a rudimentary meningeal sheath, formed of fibroblasts and macrophages, that later progressively thickens and becomes more compact. By about day 20, the fibroblasts are arranged as 16 to 20 loose rows that include bundles of collagen, oriented along the rostro-caudal axis of the cord. Even after 144 days, the meninx, although substantially thicker than normal because of the numerous fibroblast rows (20 to 30), still lacks the melanocyte layer. In cases in which the meninx at the transection site was mechanically and pharmacologically (6-hydroxydopamine) disrupted, bridge formation was essentially unchanged, and axonal regrowth continued; some regrowing axons, however, extruded from the denuded cord. Accordingly, our findings indicate that although the meningeal sheath is not essential for cord regeneration to take place, it may well facilitate recovery by providing mechanical guidance and support to the regrowing axons.

After transection, the spinal cord of the eel Anguilla quickly regrows and reconnects, and function recovers. We describe here the changes in the central canal region that accompany this regeneration by using serial semithin plastic sections and immunohistochemistry. The progress of axonal regrowth was followed in material labeled with DiI. The canal of the uninjured cord is surrounded by four cell types: S-100-immunopositive ependymocytes, S-100- and glial fibrillary acidic protein (GFAP)-immunopositive tanycytes, vimentin-immunopositive dorsally located cells, and lateral and ventral liquor-contacting neurons, which label for either γ-aminobutyric acid (GABA) or tyrosine hydroxylase (TH). After cord transection, a new central canal forms rapidly as small groups of cells at the leading edges of the transection create flat “plates” that serve as templates for subsequent formation of the lateral and dorsal walls. Profile counts and 5-bromo-2′-deoxyuridine immunohistochemistry indicate that these cells are dividing rapidly during the first 20 days of the repair process. The newly formed canal, which bridges the transection by day 10 but is not complete until about day 20, is greatly enlarged (≤100 times) and is dominated by ependymocytes that are vimentin immunopositive, but cells expressing GABA, TH, and GFAP do not appear until days 11, 13, and 16, respectively. The proliferating ependyma do not provide a supportive scaffold for the regrowing axons, inasmuch as some have crossed the bridge before the canal has formed. However, their modified phenotype suggests a role, possibly trophic, for the central canal region following injury. J. Comp. Neurol. 458:293–306, 2003. © 2003 Wiley-Liss, Inc.

Evoked field potentials were recorded from the mesencephalic (‘optic’) tectum, cerebellar corpus, midline rhombencephalon, and spinal cord of decerebrated brown trout in response to single electrical shocks given to an optic nerve. Evoked responses were also recorded from the rhombencephalon and spinal cord following stimulation (singly and with trains) to the optic tectum and to the cerebellar corpus. The potentials recorded from the tectum in response to optic nerve stimulation were similar in form to those reported by other workers from other species of teleost. The rhombencephalic responses to optic nerve and tectal stimulation were complex and comprised presumed pre- and post-synaptic events. Cerebellar stimulation evoked no detectable responses in these brain regions, but when given prior to tectal stimulation (by up to 10 ms), tectally-evoked spinal cord responses were reduced in amplitude by as much as 85%. After cerebellar ablation, there was no difference from controls in the latency, form or amplitude of any response, even when tested with paired pulse stimulation. However, when the cerebellum was ablated, rhombencephalic and spinal responses to optic nerve and tectal stimulation were markedly enhanced (by 200–300%). These clear-cut complimentary consequences of cerebellar ablation or stimulation emphasize the gain-setting role of the cerebellum and indicate, at least in relation to optically evoked motor activity, that cerebellar regulation acts at the level of the supraspinal drive to spinal motor circuits and not within the sensory centers.

Activity-dependent Fos-like expression was investigated immunohistochemically in rainbow trout (Oncorhynchus mykiss) that had performed vibratory-evoked startle responses. We found significantly higher numbers of Fos-like-immunoreactive neurons in the reticular formation, in the octavolateral area, and in several cranial nerve motor nuclei in the brain and in the motor column of the spinal cord of startled fish than in control fish. In one fish, in which stimulation did not evoke startle responses, substantial numbers of positive cells occurred in the brain, primarily in the magnocellular octavolateral nucleus. We observed Fos-like-immunoreactive neurons in cell groups that are known to participate in the startle response (e.g., the Mauthner cell) as well as in cell groups that have been proposed but until now not shown to be involved.

Evidence from mammalian studies suggests that brain derived neurotrophic factor (BDNF) and its receptor, trk B, are upregulated in neuronal cell bodies after injury. Although fish possess neurotrophins and display rapid functional and morphological recovery after central nervous system (CNS) injury, to date few studies have examined neurotrophin expression during CNS regeneration. In this study, RT-PCR was used to investigate the effect of complete spinal cord transection on the mRNA expression of BDNF and its receptor, trk B, in the eel brain at a range of timepoints after injury. The spatial expression pattern of BDNF mRNA in the brain was also assessed before and after injury using in situ hybridization. Marked changes in BDNF and trk B mRNA levels in the eel brain were not detected during the recovery period after cord transection. In addition, the spatial expression pattern of BDNF mRNA in the eel brain appeared unchanged after injury. Our results are in contrast with the increase reported in mammals but are in line with studies examining neurotrophin expression during CNS regeneration in other anamniotic vertebrates.

The spatial expression pattern of Brain-Derived Neurotrophic Factor (BDNF) mRNA in the brain of the European eel (Anguilla anguilla) was determined using non-radioactive in situ hybridization, and mapped and compared to that of other vertebrates. Riboprobes were prepared based on a partial cDNA coding sequence for A. anguilla BDNF that was amplified using degenerate primers, cloned and sequenced. As in other animal groups, in the eel, BDNF mRNA expression was seen in the telencephalon, hypothalamus, tectum, many primary and secondary sensory centers, and cranial motor nuclei. However, in contrast to mammals, BDNF mRNA expression was observed in some brain stem nuclei, such as the reticular formation, that contain cell bodies of neurons that project down the spinal cord. We suggest that these differences might relate to the continual growth of teleost fish.

The spinal cord of the eel, Anguilla, recovers function rapidly after it has been completely transected. At transection, the excitability of central pattern generating circuits in the distal denervated segments increases to such a level that undulatory movements can occur spontaneously. When this elevated neuronal activity was reduced locally, just caudal to the transection, by chronic blockade of the NMDA receptor, the normally rapid behavioural recovery was retarded. The NMDA-treated fish overcame cordotomy more slowly than untreated animals, and axons did not extend as far into the denervated cord as in untreated counterparts, until later stages of recovery. These results suggest that central pattern generating activity may facilitate axonal growth in spinal cord regeneration.

As the target musculature they innervate grows throughout life, certain segmental motoneurons from the spinal cord of Anguilla, readily identified on the basis of their form and position, also increase in size. In doing so, they present a steadily increasing target to the spinal and supraspinal neurons that innervate them. How the afferent neurons respond was assessed by measuring features of their synaptic boutons contacting the motoneuronal perikarya, as seen with electron microscopy. About 60% of the perimeter of the perikaryal profile of each motoneuron was found to be covered with synaptic bouton profiles, a value that is independent of the size of the motoneuron. Furthermore, the distances between synaptic profiles, their contact sizes (measured as apposition length) and the number and size of the vesicles each profile contains were all found to be relatively constant and also independent of motoneuronal size. In contrast, the number of synaptic profiles contacting a motoneuron correlated well with its perikaryal size. Our findings indicate that the challenge of a growing neuronal target is met by a steady increase in the number of contacting boutons, the form and spacing of which are held relatively constant; this strategy will require continual synaptic realignment at the target.

Publisher Summary This chapter discusses the morphology and activity of fish myotomal muscles. The swimming movements of fishes are of two basic patterns: (1) steady, continuous movement in which the body is bent periodically into a propulsive wave that passes backward at a velocity that is close to the swimming speed and (2) rapid, transient movement—such as accelerations and fast turns—involving large-amplitude movements. The forward velocity of a swimming fish depends on its tail-beat frequency and on the cycle period of the segmental motor output. This can be determined by the generating circuits of the spinal cord in response to sensory and descending inputs. Steady movements are the product of a single yet highly modifiable motor program in which swimming speed is changed not only by changing the cycle length of regular contraction but also by recruiting the white muscle system for faster and stronger contractions. This new recruitment results from activation of neurons projecting to the spinal cord from brainstem structures. A quite distinct program—using separate pathways, motoneurones, and muscle fibers—is implemented for very rapid, transient movements. These two programs operate independently in many fishes, but in higher teleost, they can be switched to achieve wide ranges of speed.

We describe here a computer program which solves the climate of the southwestern U.S. on a grid of points with a spacing of 7.5′ latitude and longitude. Variables computed include mean monthly values of maximum daily temperature and total precipitation. Annual values for each of these variables also are accumulated. The estimates are made with equations derived with regression procedures. Independent variables include: elevation, slope, maximum and minimum upwind elevations, ocean temperature at the coast, and distances to several key climatic controls. These are solved from boundary conditions that include: sea-surface temperature and dominant wind directions at the seasonal extremes as well as elevation and sealevel. Solutions have been obtained for the modern climate and climate characteristic of the last glacial maximum. The boundary conditions for the latter reconstruction are based upon CLIMAP reconstructions of sea-surface temperatures.

The mesencephalic nucleus of the trigeminal nerve (mes V) in the brain of the skate (Raja oscellata) was studied by electron microscopy. Mes V neurons are large (40-80 mum diameter) and are located in the periventricular grey matter. Their perikaryal cytoplasm is rich in Golgi apparatus, small mitochondria, rough endoplasmic reticulum, polysomes and bundles of neurofilaments. A striking feature is the presence of masses of glycogen granules, at times surrounded by membrane wrappings and lysosomal bodies. Two types of conventional synaptic contacts were made onto mes V perikarya and dendrites. One had round, agranular vesicles and usually also contained dense-cored vesicles, the other had flattened, pleomorphic, agranular vesicles and usually lacked dense-cored vesicles. Additional membrane complexes consisting of a region of gap junction flanked by sites of desmosomal attachment were observed to link neighbouring mes V neurons. Somato-somatic, dendro-somatic, axo-somatic, and dendro-dendritic junctions were noted. Except for the somato-somatic union, one or more chemical synapses were located close to the sites of gap junctions.

The grey matter of the spinal cord consists of two lateral, wing-like expansions in cyclostomes, whereas in chondrichthyans it is differentiated into distinct dorsal and ventral horns, an arrangement similar to that found in all other vertebrates. The spinal grey matter in cartilaginous fishes contains cells of a wide size range, although in adults the larger elements are mainly confined to the ventral horns. Myelinated fibres are absent from the cyclostomes but the white matter of the chondrichthyan cord comprises numerous well-myelinated fibres.

The statoacoustic nerve of the dogfish, Scyliorhinus canicula, was transected medial to the ganglion for the purpose of elucidating its central pathways and terminal fields. Following two to six weeks postoperative survival times, transverse, horizontal, and sagittal sections of the brain stem were stained by the Fink-Heimer silver-impregnation method to reveal degenerating axons and terminals. Fragmented axons enter the medulla and give rise to medial, descending, and ascending pathway. Fibers of the medial pathway terminate about the soma and lateral dendrites of the large cells that comprise nucleus magnocellularis; descending and ascending fibers terminate on the dendrites of the cells of ventral and superior nuclei respectively. In addition, fibers emanate from fascicles of the descending pathway to form a large field of degenerating axons and terminals within the ventromedial part of the medulla, and a substantial proportion of the fibers of the ascending pathway continues beyond the superior nucleus to terminate among the granule cells of the medial part of the vestibulolateral lobe of the cerebellum. No fragmented axons are traceable to the lateral part (auricles) of the vestibulolateral lobe, cerebellar nucleus or corpus, or those nuclei associated with the lateral-line lobes. It appears therefore that octavus terminal fields are separate from those of the lateral line at both cerebellar and medullary levels, at least at the level of the first-order neuron.

The brain stem (truncus cerebri) comprises the rhombencephalon and the mesencephalon; caudally it grades into the spinal cord while rostrally it borders the diencephalon. In all vertebrates the brain stem harbours the centres of origin and termination of all cranial nerves, except for cranial nerve I. In cartilaginous fish the brain stem contains a fairly well developed reticular formation and a number of relay centres and their associated ascending and descending connections. The cerebellum, although ontogenetically and phylogenetically a derivative of the rostral part of the rhombencephalon, is not included in the brain stem and will be considered separately in Chap. 7. However, the cerebellum and the brain stem are strongly interconnected and many of the cerebellar afferent systems originate, and almost all cerebellar efferent fibres terminate, within the brain stem.

In the majority of cartilaginous fishes the tectum mesencephali is strongly developed and differentiated into two bilateral lobes, which surround expansions of the ventricular cavity. In the species studied this cavity is most spacious in Squalus and Hydrolagus, but much reduced in Raja.