Elasmobranch Brain Research Articles

Immediate early gene expression related to learning and retention of a visual discrimination task in bamboo sharks (Chiloscyllium griseum).

Using the expression of the immediate early gene (IEG) egr-1 as a neuronal activity marker, brain regions potentially involved in learning and long-term memory functions in the grey bamboo shark were assessed with respect to selected visual discrimination abilities. Immunocytochemistry revealed a significant up-regulation of egr-1 expression levels in a small region of the telencephalon of all trained sharks (i.e., 'early' and 'late learners', 'recallers') when compared to three control groups (i.e., 'controls', 'undisturbed swimmers', 'constant movers'). There was also a well-defined difference in egr-1 expression patterns between the three control groups. Additionally, some staining was observed in diencephalic and mesencephalic sections; however, staining here was weak and occurred only irregularly within and between groups. Therefore, it could have either resulted from unintentional cognitive or non-cognitive inducements (i.e., relating to the mental processes of perception, learning, memory, and judgment, as contrasted with emotional and volitional processes) rather than being a training effect. Present findings emphasize a relationship between the training conditions and the corresponding egr-1 expression levels found in the telencephalon of Chiloscyllium griseum.Results suggest important similarities in the neuronal plasticity and activity-dependent IEG expression of the elasmobranch brain with other vertebrate groups. The presence of the egr-1 gene seems to be evolutionarily conserved and may therefore be particularly useful for identifying functional neural responses within this group.

Distribution of PACAP in the Brain of the Cartilaginous Fish Torpedo Marmorata

In this article, we investigated the distribution of pituitary adenylate cyclase-activating polypeptide (PACAP) and its mRNA by immunohistochemistry, in situ hybridization, and RT-PCR techniques, in the central nervous system of the elasmobranch Torpedo marmorata. RT-PCR analysis showed that the CNS of T. marmorata expresses a messenger encoding PACAP. The immunohistochemistry and in situ hybridization patterns were partly overlapping, with a major expression in the hypothalamo-pituitary region and, surprisingly, in the saccus vasculosus. Our results show that, in T. marmorata, PACAP is synthesized and widely distributed in the CNS, suggesting an as yet unidentified role for this peptide in elasmobranch brain physiology.

A single and novel natriuretic peptide is expressed in the heart and brain of the most primitive vertebrate, the hagfish (Eptatretus burgeri)

In teleost fish and tetrapods, the natriuretic peptide (NP) family consists of ANP (atrial natriuretic peptide), BNP (brain natriuretic peptide) and VNP (ventricular natriuretic peptide) that are secreted from the heart, and C-type natriuretic peptide (CNP) that is found in the brain. However, CNP is the only NP identified in the heart and brain of elasmobranchs, suggesting that it is the ancestral type of the NP family and that ANP, BNP and VNP appeared later in the vertebrate phylogeny. To delineate more clearly the molecular evolution of this hormone family, we determined the sequence of NP molecule(s) in evolutionarily the oldest vertebrate group, the cyclostomes. We have cloned a novel NP cDNA from the heart and brain of hagfish, Eptatretus burgeri, using the RACE method and degenerate primers that amplify all known types of NP cDNAs. The novel NP, named EbuNP after the scientific name of this hagfish, appears to be the only NP in the heart and brain, as no other NP cDNAs were amplified even after specific removal of the cloned EbuNP mRNA from the mRNA pool, except for a minor alternatively spliced EbuNP cDNA with a truncated 3'-untranslated sequence. The EbuNP was equally similar to known NPs but was not considered to be a CNP because of the presence of a C-terminal tail sequence. The EbuNP gene was abundantly expressed in the cardiac atrium, ventricle, portal heart and brain but scarcely in the intestine; no expression was observed in the gill and kidney. Mass spectrometry of affinity-purified EbuNP in plasma, heart and brain revealed a 68 amino acid peptide circulating in the blood and stored in the heart, which is cleaved at the typical cleavage signal of a processing enzyme, furin, as observed in mammalian BNP. The C-terminal Gly residue was used for amidation as is the case in eel ANP. The immunoreactive EbuNP was not detected in the brain, suggesting the presence of a different processing form in the brain. These results show that the molecular evolution of the NP family in vertebrates is more complex than previously thought.

Marked Distributional Difference of α-Melanocyte-Stimulating Hormone (α-MSH)-like Immunoreactivity in the Brain between Two Elasmobranchs (Scyliorhinus torazame and Etmopterus brachyurus): An Immunohistochemical Study

Application of α-MSH immunohistochemistry to the brain of elasmobranchs (Scyliorhinus torazame and Etmopterus brachyurus) demonstrated a marked species difference concerning the distribution of the α-MSH-like molecule in the brain. In S. torazame, α-MSH-like immunoreactive cells were present in the hypothalamus, mainly in the tuberculum posterius and the nucleus lateralis tuberis, and also in the distal and neurointermediate lobes of the hypophysis. Labeled varicose fibers were densely distributed in the hypothalamus, but they were sparse or absent in other portions of the brain. In striking contrast to the results for S. torazame, the immunoreactivity in the E. brachyurus brain was associated exclusively with the glial system, represented by astrocytes and tanycytes, throughout the central nervous system; no immunoreactivity was found in the neuronal elements. In the E. brachyurus hypophysis, the labeled cells were present in the distal and intermediate lobes, similarly to their presence in S. torazame, but in the intermediate lobe the immunoreactivity was confined to the peripheral cell cord closely adjacent to the neural lobe. The present findings are the first as regards the occurrence of α-MSH-like immunoreactivity in the glial system of the central nervous system of vertebrates and suggest diversity of expression and/or processing of proopiomelanocortin in the brain of elasmobranchs.

Differential Distribution of Gonadotropin-Releasing Hormone-Immunoreactive Neurons in the Stingray Brain: Functional and Evolutionary Considerations

Gonadotropin-releasing hormone (GnRH) is a neuropeptide that occurs in multiple structural forms among vertebrate species. Bony fishes, amphibians, reptiles, birds, and mammals express different forms of GnRH in the forebrain and endocrine regions of the hypothalamus which regulate the release of reproductive gonadotropins from the pituitary. In contrast, previous studies on bony fishes and tetrapods have localized the chicken GnRH-II (cGnRH-II) nucleus in the midbrain tegmentum and, combined with cladistic analyses, indicate that cGnRH-II is the most conserved form throughout vertebrate evolution. However, in elasmobranch fishes, the neuroanatomical distribution of cGnRH-II and dogfish GnRH (dfGnRH) cells and their relative projections in the brain are unknown. We used high-performance liquid chromatography and radioimmunoassay to test for differential distributions of various GnRH forms in tissues from the terminal nerve (TN) ganglia, preoptic area, and midbrain of the Atlantic stingray, Dasyatis sabina. These experiments identified major peaks that coelute with cGnRH-II and dfGnRH, minor peaks that coelute with lamprey GnRH-III (lGnRH-III), and unknown forms. Immunocytochemistry experiments on brain sections show that dfGnRH-immunoreactive (-ir) cell bodies are localized in the TN ganglia, the caudal ventral telencephalon, and the preoptic area. Axons of these cells project to regions of the hypothalamus and pituitary, diencephalic centers of sensory and behavioral integration, and the midbrain. A large, discrete, bilateral column of cGnRH-II-ir neurons in the midbrain tegmentum has sparse axonal projections to the hypothalamus and regions of the pituitary but numerous projections to sensory processing centers in the midbrain and hindbrain. Immunocytochemical and chromatographic data are consistent with the presence of lGnRH-III and other GnRH forms in the TN that differ from dfGnRH and cGnRH-II. This is the first study that shows differential distribution of cGnRH-II and dfGnRH in the elasmobranch brain and supports the hypothesis of divergent function of GnRH variants related to gonadotropin control and neuromodulation of sensory function.

The localization of pituitary adenylate cyclase- activating polypeptide (PACAP)-like immunoreactivity in the hypothalamo–pituitary region of an elasmobranch, stingray, Dasyatis akajei

Pituitary adenylate cyclase activating polypeptide (PACAP), a member of the secretin/glucagon/vasoactive intestinal polypeptide family of peptides, exists as 38-residue (PACAP 38) and truncated 27-residue (PACAP 27) forms, which play various roles in mammals. Recently, we isolated and characterized PACAPs with sequences very similar to that of tetrapod PACAP from the brains of two teleosts, the blue-spotted stargazer Gnathagnus elongatus and the stargazer Uranoscopus japonicus, and located PACAP-like immunoreactivities in the preoptic area, neurohypophysis and medulla oblongata of both species. In this study, PACAP-like immunoreactivity in the brain of an elasmobranch, the stingray Dasyatis akajei, was examined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) and Western blotting analysis and its distribution in the hypothalamo-pituitary region was studied immunohistochemically using the peroxidase–antiperoxidase method. The anti-PACAP 27 serum reacted with a single band of whole brain extract with a molecular weight of ca 5000. PACAP-like immunoreactive (LI) neuronal cell bodies were found in the nucleus medius hypothalami and PACAP-LI nerve fibers and terminals were seen from the nucleus to the caudal floor of the infundibular region. These results suggest that PACAP-like peptide may be present and function as a regulatory factor in the hypothalamo-pituitary region of the elasmobranch brain.

Purification and primary structure of pituitary adenylate cyclase activating polypeptide (PACAP) from the brain of an elasmobranch, stingray, Dasyatis akajei

Pituitary adenylate cyclase activating polypeptide (PACAP) was isolated from ovine hypothalami and found to exist as two amidated forms with 38 (PACAP 38) and 27 (PACAP 27) residues. The amino acid sequences of PACAPs isolated from the vertebrates, such as a bird, a frog and teleost fish, appear to be well conserved. In the present study, we attempted to isolate PACAP from the brain of an elasmobranch fish, Dasyatis akajei (stingray), which belongs to the Chondrichthyes (cartilaginous fish), by extraction of the acetone-dried powder with acetic acid, followed by successive high-performance liquid chromatography (HPLC) on a gel-filtration, a cation-exchange and two reverse-phase columns. Purification was monitored by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) and Western blotting analysis using an anti-PACAP 27 serum. The PACAP thus obtained consisted of 44 residues. The amino acid sequence of the comparable portion of its N-terminal 38 residues showed 92%, 89%, 89%, and 82% identity with those of mammalian, chicken, frog and teleost PACAPs with 38 residues, respectively. The extra six C-terminal residues of the stingray resembled those of tetrapod and teleost PACAP precursors which were deduced from the respective cDNAs. These results indicate that PACAP, which has an amino acid sequence showing high similarity with those of tetrapod and teleost PACAPs, is present in the elasmobranch brain.

Macrophage‐lymphocyte cell clusters in the hypothalamic ventricle of some elasmobranch fish: Ultrastructural analysis and possible functional significance

Previous studies have demonstrated the existence of lympho-haemopoietic tissue in the meninges and choroid plexuses of various primitive vertebrates, including the stingray Dasyatis akajei and in early human embryos. In the present study, we extend these results analyzing macrophage-lymphocyte cell clusters found in the floor of the hypothalamic ventricle of several specimens of elasmobranchs. After aseptical isolation of the brain from several specimens of smooth dogfish Triakis scyllia, cloudy dogfish Scyliorhinus torazame, gummy shark Mustelus manazo, and stingray Dasyatis akajei their hypothalamic regions were processed routinely by light, scanning, and transmission electron microscopy. The study of serial histological sections demonstrated that the macrophage-lymphocyte cell clusters proceeded from the meningeal lymphohaemopoietic tissue, reaching the ventricular lumen along large blood vessels. In this tissue, macrophages, different sized lymphocytes, lymphoblasts, granulocytes, monocytes, and developing and mature plasma cells were closely packed among a meshwork of fibroblastic reticular cell processes. It never invaded the brain parenchyma. A cell layer of glial elements and a continuous basement membrane interposed between the lymphoid tissue and the neural elements although some macrophages had migrated across the ependymal cell layer. In the ventricular lumen very irregular macrophages with long cell processes and containing abundant engulfed material of unknown origin formed big cell clusters with neighboring lymphocytes, lymphoblasts, and plasma cells, similar to those described during the immune response. Moreover, electron lucent cells which resembled the antigen-presenting cells of higher vertebrates established intimate surface cell contacts with the surrounding lymphocytes. In the third ventricle of several specimens of gummy shark, Mustelus manazo, morphologically similar cell clusters appeared but these were not connected to the meningeal lympho-haemopoietic tissue. No intraventricular cell aggregates were found in the stingray brain. Although we cannot rule out that these macrophage-lymphocyte cell clusters represent a permanent structure in the elasmobranch brain they rather seem to be only established after specific stimulation for preventing the entrance of noxious, foreign materials into the elasmobranch brain parenchyma.

Gonadotropin-releasing hormone in elasmobranch (electric ray, Torpedo marmorata) brain and plasma: Chromatographic and immunological evidence for chicken GnRH II and novel molecular forms

Gonadotropin-releasing hormone (GnRH) peptides in the brain, testis and plasma of an electric ray ( Torpedo marmorata) were investigated by gel filtration chromatography, reverse phase high performance liquid chromatography and radioimmunoassay with region-specific antisera. In the brain, two major forms of GnRH were demonstrated. One form had identical chromatographic and immunological properties to chicken GnRH II, and the second, novel, molecular form had structural features in common with mammalian, chicken II and salmon GnRHs. A minor, early-eluting immunoreactive peak, possibly also a novel GnRH, was also evident. Immunoreactive GnRH was not detected in the testis. In the plasma, a single major early-eluting immunoreactive peak was demonstrated. This peak, identical to the minor peak observed in the brain, is likely to represent a novel form of GnRH which has immunological properties in common with mammalian, chicken II and salmon GnRHs. Immunoreactive GnRH was not detected in the plasma of species from other vertebrate classes, including rabbit, chicken, monitor lizard, clawed toad, frog, cichlid fish and lamprey. The finding of chicken GnRH II in a species of Chondrichthyes adds further support to our hypothesis that this widespread structural variant may represent an early-evolved and conserved form of GnRH. The presence of a GnRH molecular form in the plasma of the electric ray suggests that GnRH may reach target organs (pituitary and gonads) via the general circulation in some species of Chondrichthyes.

Localization of corticotropin-releasing hormone (CRF)-like immunoreactivity in the central nervous system of the elasmobranch fish, Scyliorhinus canicula

The occurrence and localization of immunoreactive corticotropin-releasing factor (CRF) in the brain and pituitary of the elasmobranch fish Scyliorhinus canicula, were studied by means of specific radioimmunoassay and immunohistochemistry using the indirect immunofluorescence method. Brain and pituitary extracts showed a good cross-reactivity with the ovine CRF antiserum, but serial dilutions of tissue samples did not completely parallel the standard curve. Relatively high concentrations of CRF-like material were found within the pituitary, diencephalon, and telencephalon. CRF-like immunoreactive perikarya were observed in the preoptic nucleus and in the nucleus lateralis tuberis. Numerous immunoreactive cells appeared to be of the CSF-contacting type. CRF-like immunopositive fibers were seen to run through the hypothalamus within the ventro-medial floor of the infundibular region. A dense plexus of immunoreactive nerve endings terminated in the median eminence and the neurointermediate lobe of the pituitary. These results indicate that a neurosecretory system containing CRF-like immunoreactivity exists in the brain of elasmobranchs, a group of vertebrates which has diverged early from the evolutionary line leading to mammals. In addition, our data support the notion that a CRF-like molecule is involved in the regulation of corticotropic and melanotropic cell activity in this primitive species of fish.

Physiology of lateral line mechanoreceptive regions in the elasmobranch brain

The physiology of mechanoreceptive lateral line areas was investigated in the thornback guitarfish, Platyrhinoidis triseriata, from medulla to telecephalon, using averaged evoked potentials (AEPs) and unit responses as windows to brain functions. Responses were analysed with respect to frequency sensitivity, intensity functions, influence of stimulus repetition rate, response latency, receptive field (RF) organization and multimodal interaction. 1. Following a quasi-natural vibrating sphere stimulus, neural responses were recorded in the medullary medial octavolateralis nucleus (MON), the dorsal (DMN) and anterior (AN) nucleus of the mesencephalic nuclear complex, the diencephalic lateral tuberal nucleus (LTN), and a telencephalic area which may correspond to the medial pallium (Figs. 2, 3, 13, 14, 15, 16). 2. Within the test range of 6.5-200 Hz all lateral line areas investigated responded to minute water vibrations. Best frequencies (in terms of displacement) were between 75 and 200 Hz with threshold values for AEPs as low as 0.005 microns peak-to-peak (p-p) water displacement calculated at the skin surface (Fig. 6). 3. AEP-responses to a vibrating sphere stimulus recorded in the MON are tonic or phasic-tonic, i.e., responses are strongest at stimulus onset but last for the whole stimulus duration in form of a frequency following response (Fig. 3). DMN and AN responses are phasic or phasic-tonic. Units recorded in the MON are phase coupled to the stimulus, those recorded in the DMN, AN or LTN are usually not (Figs. 5, 8, 9). Diencephalic LTN and telencephalic lateral line responses (AEPs) often are purely phasic. However, in the diencephalic LTN tonic and/or off-responses can be recorded (Fig. 11). 4. For the frequencies 25, 50, and 100 Hz, the dynamic intensity range of lateral line areas varies from 12.8 to at least 91.6 dB (AEP) respectively 8.9 and 92 dB (few unit and single unit recordings) (Fig. 7). 5. Mesencephalic, diencephalic, and telecephalic RFs, based on the evaluation of AEPs or multiunit activity (MUA), are usually contralateral (AN and LTN) or ipsi- and contralateral (telencephalon) and often complex (Figs. 10, 12, 16). 6. In many cases no obvious interactions between different modalities (vibrating sphere, electric field stimulus, and/or a light flash) were seen. However, some recording sites in the mesencephalic AN and the diencephalic LTN showed bimodal interactions in that an electric field stimulus decreased or increased the amplitude of a lateral line response and vice versa (Fig. 13 B).

Acid-induced injury in elasmobranch brain

Sodium lactate injection into rat brain produces coagulation necrosis consistent with infarction when pH o is held at ⩽5.30 for 20 min. Such injury may result from excessive astroglial acidification. If true, then brain damage from acidosis in elasmobranchs might evolve differently since glial reaction there to another necrotizing injury, exposure to extreme cold, is dissimilar from that seen in mammals. Accordingly, pH o was monitored and sodium lactate (pH 4.00–7.00) injected into skate ( Raja erinacea) cerebella. Necrosis was seen only when pH o was ⩽4.86 for 20 min; and pH o rise after injections was unaffected by those which destroyed brain, and not slowed as in rat. Thus elasmobranchs are less susceptible to irreversible brain injury from acidosis, a capacity which may result from their lower body temperature compared to mammals.

Glutamine synthetase isozymes in elasmobranch brain and liver tissues.

Glutamine synthetase is present as isozymic forms in the elasmobranchs Squalus acanthias (dogfish shark) and Dasyatis sabina (stingray). Subcellular fractionation of elasmobranch brain and liver tissue shows the enzyme to be predominantly cytosolic in the former tissue and mitochondrial in the latter. For the cytosolic brain enzyme, the subunit Mr equals 42,000 in the stingray and 45,000 in the shark, as determined by sodium dodecyl sulfate-gel electrophoresis/Western blotting. The subunit Mr = 45,000 and 47,000, respectively, for stingray and dogfish mitochondrial liver enzymes. Translation of total brain RNA from both species gives immunoprecipitable nascent peptides of the same size as their respective mature enzymes. However, in liver tissue, translation of glutamine synthetase mRNA yields peptides of higher Mr than that of the mature enzymes. In dogfish liver, Mr = 50,000 for the translation product and, in stingray liver, Mr = 48,000. This suggests that the translocation of the enzyme into liver mitochondria may be via a signal or leader sequence mechanism. The larger liver isozyme of elasmobranch glutamine synthetase is found in kidney where it is also known to be mitochondrial. The smaller cytosolic isozyme occurs in retina, heart, gill, and rectal gland tissue as well as in brain.

Elasmobranch oculomotor organization: Anatomical and theoretical aspects of the phylogenetic development of vestibulo‐oculomotor connectivity

The oculomotor organization of two elasmobranch species, smooth dogfish (Mustelus canis) and little skate (Raja erinacea), was studied by investigating the extraocular muscle apparatus and the oculomotor motoneuron distribution. The macroscopic appearance of the eye muscles was similar to any lateral-eyed vertebrate species (e.g., goldfish, rabbit). The size of extraocular muscles was expressed by counting single muscle fibers and comparing cross-sectional areas of the extraocular muscles. There were significant differences in the number of fibers in the six extraocular muscles in dogfish, but not in skate. Fiber sizes varied considerably; thus, the number of fibers did not relate to cross-sectional areas. In the dogfish, no one pair of agonist-antagonist extraocular muscles was larger than the others, suggesting that there was no preference for eye movements in a particular plane of space. However, the lateral rectus was more than twice the size of most of the other muscles. In the skate, cross-sectional areas of the horizontal eye muscles were smaller than those of the vertical eye movers. This may indicate a reduced utilization of horizontal eye muscles, which may reflect the bottom-dwelling habitat and mode of locomotion of the skate. The distribution of the extraocular motoneurons was determined by injecting horseradish peroxidase (HRP) into single eye muscles. Medial rectus, superior rectus, and superior oblique motoneuron populations were located contralateral to their respective muscles. Lateral rectus, inferior rectus, and inferior oblique motoneurons were located ipsilateral to their muscles. This distribution is in contrast to almost all other vertebrates studied thus far, where medial rectus motoneurons are located ipsilateral to the muscle which they innervate. The oculomotor arrangement in elasmobranchs is likely to have consequences for the circuitry responsible for the production of conjugate compensatory eye movements in the horizontal plane. We hypothesize that, in contrast to other vertebrates, the basic elasmobranch vestibulo-ocular reflex pathway consists of three identically structured three-neuron-arcs connecting the three semicircular canals to their respective extraocular muscles. This innervation pattern may constitute a special feature of the elasmobranch brain or a phylogenetically older arrangement of eye movement pathways.

Reconstitution of ciliary membranes containing tubulin.

Membranes from the gill cilia of the mollusc Aequipecten irradians may be solubilized readily with Nonidet P-40. When the detergent is removed from the solution by adsorption to polystyrene beads, the proteins of the extract remain soluble. However, when the solution is frozen and thawed, nearly all of the proteins reassociate to form membrane vesicles, recruiting lipids from the medium. The membranes equilibrate as a narrow band (d = 1.167 g/cm3) upon sucrose density gradient centrifugation. The lipid composition of reconstituted membranes (1:2 cholesterol:phospholipids) closely resembles that of the original extract, as does the protein content (45%). Ciliary calmodulin is the major extract protein that does not associate with the reconstituted membrane, even in the presence of 1 mM calcium ions, suggesting that it is a soluble matrix component. The major protein of reconstituted vesicles is membrane tubulin, shown previously to differ hydrophobically from axonemal tubulin. The tubulin is tightly associated with the membrane since extraction with 1 mM iodide or thiocyanate leaves a vesicle fraction whose protein composition and bouyant density are unchanged. Subjecting the detergent-free membrane extract to a freeze-thaw cycle in the presence of elasmobranch brain tubulin or forming membranes by warming the extract in the presence of polymerization-competent tubulin yields a membrane fraction with little incorporated brain tubulin. This suggests that ciliary membrane tubulin specifically associates with lipids, whereas brain tubulin preferentially forms microtubules.

Fine structure and permeability of capillaries in the stria vascularis and spiral ligament of the inner ear of the guinea pig.

The blood capillaries in the stria vascularis and the spiral ligaments of guinea pigs were studied by electron microscopy with freeze-fracture and thin section methods, including tracer experiments with horseradish peroxidase (HRP) and microperoxidase (MP). The endothelial cells of the capillaries of both tissues are connected by tight junctions, and contain about the same number of micropinocytotic vesicles. In cases of intravascular administration before fixation, both of the tracers stained the perivascular space and almost all endothelial vesicles in the stria vascularis. On the other hand, the perivascular space and many vesicles in the spinal ligament were unstained. The endothelial tight junctions in the stria vascularis prevented the penetration of HRP, but sometimes allowed the penetration of MP. Those of the spiral ligament were impermeable to both tracers. In cases of tracer administration after fixation, leakage spots of HRP from capillaries were sparsely located all over the stria vascularis. Transendothelial channels and isolated fenestrae formed by micropinocytotic vesicles were detected. It is concluded that the capillaries of the stria vascularis are similar to the muscle capillaries and to the capillaries of the elasmobranch brain, whereas those in the spiral ligament are similar to the brain capillaries of higher vertebrates.

Auditory centers in the elasmobranch brain stem: Deoxyglucose autoradiography and evoked potential recording

Elasmobranchs are sensitive to low frequency sound, and electrophysiological studies have demonstrated acoustic responses from the ear. Five primary projections from the ear to the medulla have been found, but individually they could not be identified with either the auditory or the equilibrium modality since they originate in a mixed nerve. Metabolic mapping in the brain of the thornback guitarfish with [14C]2-deoxyglucose autoradiography and acoustic stimulation provided tentative identifications of acoustic centers in cell plate X of the medial octavolateralis nucleus, the anterior octaval nucleus, the nucleus of the lateral lemniscus, and the ventromedial division of the lateral mesencephalic nucleus. Evoked potential recordings confirmed acoustic activity in those sites and additionally in the reticular formation, and the lateral granule cell mass of the auricle. In tests in the mesencephalon the evoked potential disappeared within the range of elasmobranch behavioral thresholds and when the eighth nerves were cut, but was not changed when all 4 lateral line nerves were cut. The identified acoustic centers resemble those found in auditory lemniscal pathways in mammals and other tetrapods, but the most recent ancestor common to elasmobranchs and tetrapods lived 400 million years ago. Therefore, a basic auditory lemniscal pathway may be a longstanding feature of the vertebrate brain.

ON THE COMPOSITION AND STRUCTURE OF INDIVIDUAL GANGLIOSIDES FROM THE BRAIN OF ELASMOBRANCHES

AbstractGangliosides with a short carbohydrate chain: II3(NeuAc)2‐LacCer, II3(NeuAc)2‐GgOse3Cer, II3(NeuAc)3‐LacCer, II3NeuAc‐LacCer, and II3NeuAC‐GgOse3 Cer, were found to be predominant in the brain of 8 species of cartilaginous fish, elasmobranches. N‐acetylneuraminic acid was the only sialic acid found in these gangliosides, the N‐glycolyl derivative being practically absent. 4‐Sphingenine was shown to be the predominant sphingoid in elasmobranch brain gangliosides.The sequential enzymatic hydrolysis of II3(NeuAc)2 ‐LacCer from shark and ray brain by acylneur‐aminyl hydrolase (EC 3.2.1.18) and β‐D‐galactoside‐galactohydrolase (EC 3.2.1.23), as well as permethylation studies, provide further evidence for the following structure of this major elasmobranch brain ganglioside:

Comparative studies on synaptosomes: Applicability of the rapid method for preparing synaptosomes to elasmobranch brain

A flotation technique for the rapid preparation of synaptosomes, originally developed for invertebrate nervous tissue, has now been successfully applied to that of an elasmobranch fish (Mustelis canis, dogfish). The technique involves submitting the supernatant, obtained after a homogenate has been centrifuged at low speed to remove nuclei and tissue debris to centrifugal fields of intermediate intensity (10(6) g/min), appears to separate well-sealed synaptosomes from those less well sealed as judged by the criteria of osmotic shrinkage, enzyme occlusion, and choline uptake. The sealed synaptosomes do not equilibrate with the 0.8 M sucrose used as the homogenization medium and rise to form a coherent pellicle at the top of the tube. Due to the short (1-1/1/2 hr) preparation time, such synaptosomes may well prove useful in further metabolic studies.

Phospholipids and glycolipids in the brain of marine fish

1. 1. The brain phospholipid content of most of the marine fishes studied varies from 1000 to 2000 μg of P/g of wet wt. The brain phospholipid patterns of the teleosis studied are similar while the elasmobranch brain contains relatively less phosphatidyl choline and more phosphatidyl ethanolamine than the teleost one. 2. 2. Fatty acid composition of phosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl serine, phosphatidyl inositol and sphingomyelin has been studied. Each of the phospholipids studied has a particular fatty acid pattern resembling that of the phospholipids from the brain of warm-blooded animals. All the fish brain phospholipids studied are less saturated and contain more polyunsaturated fatty acids of the ω3 family than the phospholipids from the brain of warm-blooded vertebrates. The phospholipid fatty acid composition in the brain of the only abyssal bottom fish available ( Leucicorus sp.) differs very much from that of other fishes studied. 3. 3. Cerebroside content in the brain of teleosts studied varies from 1·1 to 4·9 mg/g of wet wt, the sulfatide content being 0·9–1·9 mg/g, while in shark brain the cerebroside and sulfatide concentration is considerably higher. The fish brain unlike the warm-blooded brain is characterized by the predominance of cerebrosides with normal fatty acids over oxycerebrosides. 4. 4. Fish brain ganglioside content varies from 150–515 μg of sialic acid/g of wet wt having its highest concentration in elasmobranch brain. Considerable amounts of tetra and pentasialogangliosides were found in the fish brain studied, their concentration being especially high in the teleost brain. Fatty acid composition of fish brain gangliosides was studied as well.