On the organization of receptive fields of retinal spot detectors projecting to the fish tectum: Analogies with the local edge detectors in frogs and mammals.

Responses of ON- and OFF-ganglion cells (GCs) were recorded extracellularly from their axon terminals in the medial sublamina of tectal retino-recipient layer of immobilized cyprinid fish (goldfish and carp). These units were recorded deeper than direction selective (DS) ones and at the same depth where responses of orientation selective (OS) GCs were recorded. Prominent responses of these units are evoked by small contrast spots flickering within or moving across their visual field. They are not selective either to the direction of motion or to the orientation of stimuli and are not characterized by any spontaneous spike activity. We refer to these fish GCs as spot detectors (SDs) by analogy with the frog SD. Receptive fields (RFs) of SDs are organized concentrically: the excitatory center (about 4.5°) is surrounded by opponent periphery. Study of interactions in the RF has shown that inhibitory influences are generated already inside the central RF area. This fact suggests that RFs of SDs cannot be defined as homogeneous sensory zone driven by a linear mechanism of response generation. Physiological properties of fish SDs are compared with the properties of frog SDs and analogous mammalian retinal GCs-local edge detectors (LEDs). The potential role of the SDs in visually guided fish behavior is discussed.

A GABAergic tecto-tegmento-tectal pathway in pigeons.

Previous studies have demonstrated that the optic tecta of the left and right brain halves reciprocally inhibit each other in birds. In mammals, the superior colliculus receives inhibitory γ-aminobutyric acid (GABA)ergic input from the basal ganglia via both the ipsilateral and the contralateral substantia nigra pars reticulata (SNr). This contralateral SNr projection is important in intertectal inhibition. Because the basal ganglia are evolutionarily conserved, the tectal projections of the SNr may show a similar pattern in birds. Therefore, the SNr could be a relay station in an indirect tecto-tectal pathway constituting the neuronal substrate for the tecto-tectal inhibition. To test this hypothesis, we performed bilateral anterograde and retrograde tectal tracing combined with GABA immunohistochemistry in pigeons. Suprisingly, the SNr has only ipsilateral projections to the optic tectum, and these are non-GABAergic. Inhibitory GABAergic input to the contralateral optic tectum arises instead from a nearby tegmental region that receives input from the ipsilateral optic tectum. Thus, a disynaptic pathway exists that possibly constitutes the anatomical substrate for the inhibitory tecto-tectal interaction. This pathway likely plays an important role in attentional switches between the laterally placed eyes of birds. J. Comp. Neurol. 524:2886-2913, 2016. © 2016 Wiley Periodicals, Inc.

Two parallel ascending pathways from the dorsal octavolateral nucleus to the midbrain in the paddlefish Polyodon spathula.

The paddlefish is a passive electrosensory ray-finned fish with a special rostral appendage that is covered with thousands of electroreceptors, which makes the fish extremely sensitive to electric fields produced by its primary prey, small water fleas. We reexamined the electrosensory pathways from the periphery to the midbrain by injecting the neuronal tracer BDA into different branches of the lateral line nerve and into different parts of the dorsal octavolateral nucleus (DON) and the tectum. Primary afferents from the anterior to posterior body axis terminate in different areas in the mediolateral axis of the DON, the first electrosensory processing station. Previous studies showed that DON neurons project to the tectum and two different areas in the tegmentum. Now, we have found differences between the anterior and the posterior DON. Fibers from the anterior DON project unilaterally to the contralateral tectum while its posterior neurons project bilaterally to two nuclei in the tegmentum, the torus semicircularis and the lateral mesencephalic nucleus. This study is the first to show that two different populations of ascending neurons project to two different targets in the midbrain. These two pathways are likely to have different functions and further investigations may reveal the functional significance of these two parallel ascending systems.

Synaptic membrane freezing affects modulatory sites in avian central nervous system GABA(A) receptor.

Studies were carried out to determine whether barbiturates and neurosteroids share common recognition sites at the GABA(A) receptor complex in avian CNS. To achieve this, differentially prepared fresh and frozen synaptic membranes were used. Both the barbiturate, pentobarbital, and the neurosteroid, 3alpha-hydroxy-5alpha-pregnan-20-one, were able to stimulate GABA binding in both types of membranes. Stimulation differed markedly when both drugs were added jointly to different treated tissue. In frozen membranes drugs acted synergistically and were differentially displaced by picrotoxinin, while in fresh ones, where both compounds were inhibited by the convulsant, this additivity was absent. Post-freezing wash supernatants were collected and used as a source of putative endogenous factors involved in the above mentioned membrane differences. Addition of a high molecular weight fraction from supernatants to frozen synaptic membranes led to an inhibition of barbiturate and neurosteroid potentiation, as well as a loss of their additive effect. Our results indicate that GABA(A) receptor modulation by barbiturates and neurosteroids is affected by synaptic membrane treatment, with a common modulatory site in fresh membranes and separate recognition sites after a freeze-thawing procedure. There may also be endogenous factors involved in overlapping of modulatory sites, which would thus regulate GABA(A) receptor functionality by direct interaction with the complex.

Immunocytochemical mapping of NPY and VIP neuronal elements in the cat subcortical visual nuclei, with special reference to the pretectum and accessory optic system.

The aim of this study was to describe the distribution patterns of neuropeptide Y (NPY) and vasoactive intestinal polypeptide (VIP)-immunoreactive (ir) neuronal elements in subcortical visual centers of the cat. Numerous NPY-ir neurons were present in the feline nucleus of the optic tract and in the anterior pretectal nucleus. Only a few NPY-ir neurons were found in the posterior, medial and olivary pretectal nuclei and in the accessory optic nuclei. Diffuse and heavily beaded NPY-ir fiber plexuses were observed throughout the superior colliculus, pretectum, and accessory optic system. Extensively arborising NPY-ir fibers were present in the mesencephalon and ventral lateral geniculate nucleus, while the dorsal visual thalamic nuclei contained only a few NPY-ir fibers. VIP-ir cells were present mainly in the accessory optic nuclei, and they were absent in the dorsal visual thalamus. Both NPY- and VIP-ir neurons were multipolar and fusiform in shape in the regions studied. Enucleation did not alter the appearance of NPY- and VIP-containing neuronal elements in the superior colliculus and pretectum while in the thalamus a subset of NPY-ir fiber population disappeared, indicating their retinal origin. Although there is a partial overlap in the topographical localization of the NPY- and VIP-ergic neurons in the pretectum, the colocalization of the two peptides could not be demonstrated. The present observations demonstrate the existence of two different and separate peptidergic (NPY and VIP) neuronal populations in the pretectum.

Effect of a 30 kD protein from tectal extract of rat on cultured retinal neuron.

For the first time we have shown here that the constituent from tectal extract (Te) which can support and promote the survival and growth of the retinal ganglion cell is a 30 kD protein. (i) Using MTT colorimetric microassay to measure the optical density for the survival and growth of the cultured retinal neuron, it was found that the optical densities for the experimental cultures with either Te, or its 10-30 kD fraction, or its greater than or equal to 30 kD fraction were 2-4 times that of the control culture without Te (P less than 0.01). This indicated that experimental cultures were more active in growth. (ii) The retinal neurons cultured on Te Phast gels showed that large retinal neurons (greater than 18 microns) grew only on the gel region containing the 30 kD protein. The retrograde prelabelled with horseradish peroxidase (HRP) for retinal ganglion cells indicated that these surviving neurons grown on Te Phast gel were HRP positive, i.e. retinal ganglion cells. (iii) The retinal explants cultured with 30 kD gels at a close distance of 1 mm apart revealed that many neurite outgrowths, up to 400 microns, extended from the retinal explants towards 30 kD gels, and that many individual cells and tissue masses migrated out from the explants and grew onto 30 kD gels. They were stained anti-neuron specific enolase and anti-Thy 1.1 positive, indicating they are retinal ganglion cells. Therefore, we concluded that the 30 kD protein is the neuronotrophic factor in the tectal extract specific for retinal ganglion cells.

Purification and characterization of an alpha-bungarotoxin receptor that forms a functional nicotinic channel.

Neither the structure nor the function of alpha-bungarotoxin (alpha Bgtx) binding molecules in the nervous system have yet been completely defined, although it is known that some of these molecules are related to cation channels and some are not. Using an improved method of affinity chromatography, we have isolated a toxin binding molecule from chicken optic lobe that contains at least three subunits with apparent Mr values of 52,000, 57,000, and 67,000. The Mr 57,000 subunit binds alpha Bgtx and seems to be present in two copies per receptor. The receptor is recognized by antibodies raised against the alpha Bgtx receptors of human neuroblastoma cells, fetal calf muscle, and chicken optic lobe but not by antibodies raised against Torpedo acetylcholine receptor, the serum of myasthenic patients, or monoclonal antibody, 35. 125I-labeled alpha Bgtx binding to the isolated receptor is blocked, with the same potency, by nicotinic agonists and antagonists, such as nicotine, neuronal bungarotoxin and, d-tubocurarine. When reconstituted in a planar lipid bilayer, the purified alpha Bgtx receptor forms cationic channels with a conductance of 50 pS. These channels are activated in a dose-dependent manner by carbamylcholine and blocked by d-tubocurarine.

[The characteristics of the cell aggregation of the tectum mesencephali in chick embryos and of the cerebral cortex in newborn rats in mixed cultures]. / Osobennosti agregatsii kletok kryshi srednego mozga émbrionov kur i kory golovnogo mozga novorozhdennykh krys v smeshannykh kul'turakh.

To determine species-specific cells in mixed culture, obtained after simultaneous cultivation of dissociated cells of chick embryo midbrain tegmentum and newborn albino rat cerebral cortex, various DNA amounts were used for testing chick and rat cells. The study of cell structure of the aggregates revealed that in addition to the aggregates consisting of cells belonging to only one species of animals, chimera aggregates also exist made of cells of both species of animals, basically of chick embryo glial cells and newborn rat neurons.

Brain and muscle nicotinic acetylcholine receptors are different but homologous proteins.

An alpha-bungarotoxin-binding protein was purified from chick optic lobe and brain by an improved method. Previous and present observations justify its designation as a brain nicotinic acetylcholine receptor (AcChoR). It contains subunits whose apparent molecular weights are somewhat larger than those of subunits of peripheral AcChoRs. The size of the optic lobe AcChoR complex is greater than that of the peripheral receptor when estimated from its sedimentation behavior. Brain AcChoR subunits can be specifically precipitated by a monoclonal antibody directed against chick muscle AcChoR. Amino-terminal amino acid sequence analysis was performed on AcChoR preparations and isolated subunits from the optic lobe and from the rest of the chick brain. The sequences obtained demonstrate that, at least for the lowest molecular weight component, the AcChoRs from different brain areas are identical and they are highly homologous to muscle AcChoR. It is concluded that the brain alpha-bungarotoxin-binding protein is indeed a nicotinic AcChoR and is encoded by a set of genes that is different from, but strongly related to, that for the muscle AcChoR.

Nicotinic acetylcholine receptor from chick optic lobe.

An alpha-bungarotoxin-sensitive nicotinic cholinergic receptor from chick optic lobe has been completely purified. Its standard sedimentation coefficient is 9.1 S. The value near 12 S reported for the related component from other brain regions can be reproduced when the initial extraction is by Triton X-100 (rather than Lubrol PX), but other protein is then complexed with it. A single subunit of apparent molecular weight 54,000 is detected, and this subunit is specifically labeled by bromo-[3H]acetylcholine, but only after disulfide reduction. The same size subunit likewise is labeled in the protein (purified similarly) from the rest of the chick brain which can also bind alpha-bungarotoxin and nicotinic ligands. Immunological crossreactivity is demonstrated between both of these proteins with an antiserum to pure acetylcholine receptor from skeletal muscle. The acetylcholine receptor from chick optic lobe and the alpha-bungarotoxin-binding protein from the rest of the brain appear similar or identical by a series of criteria and are related to (but with differences from) peripheral acetylcholine receptors.

Biochemical studies of chick brain development and maturation: I. Alterations in the macromolecule content and cell-free protein synthesis.

Changes in macromolecule content (DNA, RNA, and protein) and cell-free protein synthetic activity in chick brain developing from 9-day-old embryos to 1 1/2-year-old adults were studied in three brain regions: cerebrum, cerebellum, and optic lobes. The accumulation of DNA, RNA, and protein in three brain regions occurred each in its own characteristic pattern although three distinct phases could be observed. The most rapid accumulation always occurred during embryonic development followed by a slower accumulation from hatching to 3-mo. with very little or no change in the brain macromolecule content thereafter. In all three brain regions, the cell-free protein synthetic activity increased during embryonic development, reaching maximum around hatching then decreased substantially (50-70%) after hatching. Comparison of 18-day-old embryonic and 1 1/2-year-old adult brain tissue showed that there was no significant alteration in the relative proportion of membrane-bound and free ribosomes. When the products of cell-free protein synthesis by embryonic and adult brain preparations were compared by SDS polyacrylamide gel electrophoresis, it was found that the percentage of various proteins synthesized by embryonic brain tissue was very similar to that of the adult. It is concluded that the decrease in cell-free protein synthesis is due to a general decrease in the synthesis of all types of proteins rather than a decrease in a specific group or groups of proteins.

An adrenergic mechanism for visual neuroregulation.

Studies of the diffuse electroretinogram (ERG) and evoked reponses in the optic lobe in the baby chick have shown that epinephrine produces changes in both. Epinephrine produces enhancement of the a-wave and depression of the b- and d-waves in both intact and isolated eyes. In the optic lobe it produces enhancement of both components of the evoked response. The presence of epinephrine in the vertebrate retina and the optic lobe of chicks suggests that these changes may reflect the involvement of epinephrine in synaptic transmission.