Regional levels of neurotransmitter markers in the pigeon telencephalon: a comparison with possibly homologous areas of the rat telencephalon.

The levels of cholinergic, gamma-aminobutyric acidergic (GABAergic), and excitatory amino acid neurotransmitter markers have been measured in 18 regions of the pigeon telencephalon as well as in supposedly homologous areas of the rat telencephalon. Among the basal telencephalic areas, some similar patterns of regional distribution were observed, with the noticeable exception of the ratio of levels of cholinergic markers between the striatum and globus pallidus, which was much larger in the rat than in the pigeon. In the rat cortical areas, some interesting differences were noticed among the archicortex, the paleocortex, and various parts of the neocortex. In particular, the area identified as prefrontal cortex by previous studies was significantly richer in cholinergic and excitatory amino acid markers and poorer in GABAergic activity than other neocortical regions. In the pigeon, presumedly neocortical equivalent areas--in particular, those constituting the dorsal ventricular ridge--were quite variable in levels of cholinergic markers, and some apparently well-established areas homologous to mammalian neocortex showed exceptionally low levels of cholinergic markers. The higher variability in levels of neurotransmitter-related markers shown by cortically equivalent areas of the avian dorsal ventricular ridge, as compared with the more uniform pattern present in basal telencephalic regions, may be the result of a greater plasticity of these structures during evolution, in response to different selective pressures.

Neurotoxic effect of 1-methyl-4-phenylpyridinium ion on dopaminergic neurons of the retina of goldfish.

Dopaminergic neurons of the goldfish retina were selectively destroyed after a single intravitreal injection of 1-methyl-4-phenylpyridinium ion (MPP+). The ultrastructural analysis of the retina 3 days after toxin administration shows darkening of some retinal neurons present in the inner nuclear layer including their cytoplasmic processes. Both uptake and release of dopamine were reduced in the toxin-injected retina, whereas choline acetyltransferase (ChAT) and glutamic acid decarboxylase (GAD) activities, as well as the uptake of D-[3H]aspartate were not affected.

Effect of AF64A on the cholinergic systems of the retina and optic tectum of goldfish.

AF64A, a presumed selective cholinergic neurotoxin has been used to study the effect on cholinergic systems of the goldfish retina and optic tectum. Toxin injection in the vitreum and in the optic tectum caused a selective decrease of choline acetyltransferase activity in both areas, while no significant decrease of glutamate decarboxylase and D-3H aspartate uptake were observed at different times after the injections. The effect was particularly dramatic in the retina of long term-injected animals, where choline acetyltransferase dropped to practically zero level. The ultrastructural analysis showed selective degeneration of some neurons in the amacrine and ganglion cell layer of the retina as well as of synaptic terminals and neuronal cell bodies in the optic tectum. The results favour a selective cholinotoxicity of AF64A in fish nerve tissue at doses substantially higher than those found to have additional unselective effects in mammals.

Evolution of neurotransmitter-related markers in the vertebrate telencephalon. Comparative microchemical study in discrete brain regions of a frog and a turtle.

1. Neurochemical markers related to cholinergic, GABAergic and glutamatergic/aspartatergic neurotransmission have been measured in telencephalic areas obtained by microdissection from frog (Rana esculenta) and turtle (Pseudemys scripta elegans) brain. 2. In both species, pallial areas showed remarkably higher levels of synaptosomal D-3H-aspartate high affinity uptake than basal regions. Conversely, striatal and septal areas possessed higher levels of the GABAergic marker glutamate decarboxylase (GAD) than the pallium. 3. A differential distribution of GAD was noticed in striatal regions, highest levels of the enzyme being present in the ventral striatum, followed by the nucleus accumbens and the dorsal striatum. 4. Cholinergic markers choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) were rather uniformly distributed in the frog telencephalon, while, in the turtle, cholinergic markers were several-fold higher in the basal telencephalon, particularly in the striatum, than in the pallium. 5. The turtle dorsal ventricular ridge possessed ChAT levels more similar to the striatal than to the cortical ones. On the contrary, D-3H-aspartate uptake in the dorsal ventricular ridge was close to the highest levels found in cortical areas. 6. The quantitative neurochemical approach adopted for the present study appears to be a useful tool to investigate the problem of homologies and to gain new information on the evolution of neuron populations and neuronal connections in the vertebrate telencephalon.

Cholinergic projections in the telencephalo-habenulo-interpeduncular system of the goldfish.

Experiments were performed in order to see whether acetylcholine is a neurotransmitter in the telencephalo-habenulo-interpeduncular system of the goldfish. After telencephalic ablation choline acetyltransferase decreased by 53% in the habenular nuclei but was unchanged in the interpeduncular nucleus. After combined telencephalic and habenular lesions, the enzyme level dropped by 95% in the interpeduncular nucleus. These results suggest the existence of a telencephalo-habenular and a habenulo-interpeduncular cholinergic projection in the goldfish, as previously demonstrated in mammals. Acetylcholinesterase histochemistry additionally demonstrates some similarities between the habenulae and the interpeduncular nucleus in the goldfish and in mammals.

Regional levels of neurotransmitter-related markers in the brain of the weakly electric fish Gnathonemus petersii.

The regional distribution of neurochemical markers for cholinergic, GABAergic and glutamatergic/aspartatergic transmission has been studied in several brain areas of the weakly electric fish Gnathonemus petersii. In particular, brain regions related to reception, relaying and processing of electrosensory modalities have been examined. The regional distribution of neurotransmitter-related markers is considered in the light of available anatomical data and is discussed with particular emphasis on differences among different parts of the hypertrophic valvula cerebelli and other cerebellar structures.

Modification of ultrastructural and neurochemical parameters in synaptosomes of the retino-deprived goldfish optic tectum.

Neurochemical parameters associated with cholinergic and excitatory amino acid transmission, were measured in synaptosomes of the goldfish optic tectum at different times after unilateral eye ablation. Significant decreases in choline acetyltransferase and acetylcholinesterase were measured 12 and 30 days after enucleation. The high affinity choline uptake did not parallel the decrease in cholinergic enzymes. Instead there was a significant increase of the uptake per unit of protein (though not relative to the total number of tectal synaptosomes). No decrease of the high affinity D-3H aspartate uptake was measured in the deafferentated optic tectum. Electron microscopic observations showed a correspondence between the time course of cholinergic enzyme decrease and the degeneration of retinal afferents to the tectum. The present results support the notion that acetylcholine is a better candidate than the excitatory amino acids for a neurotransmitter role in the fish optic tectum.

Cholinergic, GABAergic and excitatory amino acidic neurotransmission in the goldfish vagal lobe.

Some neurotransmitter systems operating in the goldfish vagal lobe, an hypertrophied gustatory center, have been studied by means of experimental (kainic acid injection and vagal rhizotomy), neurochemical and ultrastructural methods. The use of the neurotoxin, kainic acid, revealed the existence of cholinergic and GABAergic neurons in the vagal lobe. The results of histochemical observations and biochemical assays performed after rhizotomy of sensory and motor vagal roots, suggest that the motor neurons of the vagal motor layer are cholinergic. The same experiments also indicate that the primary gustatory afferents distributing to the sensory layer of the vagal lobe are, at least in part, cholinergic. By contrast, no decrease of excitatory amino acid uptake was demonstrated following the experimental lesions. GABA is likely to play an important role in the goldfish vagal lobe, particularly in the sensory layer, where the highest level of its synthetic enzyme, glutamate decarboxylase, is recorded. The significant decrease of glutamate decarboxylase in the sensory layer after vagal rhizotomy suggests that either GABAergic primary afferents reach the vagal lobe, or that deafferentation results in a decreased GABA synthesis in intrinsic GABAergic neurons.

Kainic acid differentially affects the synaptosomal release of endogenous and exogenous amino acidic neurotransmitters.

Presynaptic actions of kainic acid have been tested on uptake and release mechanisms in synaptosome-enriched preparations from rat hippocampus and goldfish brain. Kainic acid increased in a Ca2+-dependent way the basal release of endogenous glutamate and aspartate from both synaptosomal preparations, with the maximum effect (40-80%) being reached at the highest concentration tested (1 mM). In addition, kainic acid potentiated, in an additive or synergic way, the release of excitatory amino acids stimulated by high K+ concentrations. Kainic acid at 1 mM showed a completely opposite effect on the release of exogenously accumulated D-[3H]aspartate. The drug, in fact, caused a marked inhibition of both the basal and the high K+-stimulated release. Kainic acid at 0.1 mM had no clear-cut effect, whereas at 0.01 mM it caused a small stimulation of the basal release. The present results suggest that kainic acid differentially affects two neurotransmitter pools that are not readily miscible in the synaptic terminals. The release from an endogenous, possibly vesiculate, pool of excitatory amino acids is stimulated, whereas the release from an exogenously accumulated, possibly cytoplasmic and carrier-mediated, pool is inhibited or slightly stimulated, depending on the external concentration of kainic acid. Kainic acid, in addition, strongly inhibits the high-affinity uptake of L-glutamate and D-aspartate in synaptic terminals. All these effects appear specific for excitatory amino acids, making it likely that they are mediated through specific recognition sites present on the membranes of glutamatergic and aspartatergic terminals. The relevance of the present findings to the mechanism of excitotoxicity of kainic acid is discussed.

Neurotransmitter-related markers in the normal and experimentally lesioned telencephalon of the goldfish.

The telencephalon of teleost fish shows high affinity uptake for D-[3H]aspartate, intermediate levels of GABAergic markers and low levels of cholinergic enzymes. Experimental results (resection of the olfactory tracts or unilateral kainic acid administration in the telencephalon) suggest that: the projection from the olfactory bulbs to telencephalic targets is mediated by aspartate and/or glutamate; and a population of GABAergic neurons is present in a telencephalic area which is considered homologous to part of the striatal complex of land vertebrates. From the present results, it appears that the neurochemical approach can be used, in conjunction with neuroanatomical methods, to study evolutionary problems of telencephalic function.

Effect of kainic acid, glutamate, and aspartate on CO2 production by goldfish tectal slices.

For a study of the excitatory effect of kainate, glutamate, and aspartate in the goldfish optic tectum, these substances were tested on the production of CO2 from radioactive glucose in tectal slices incubated in Krebs-Ringer medium for fish. Kainate increased the rate of CO2 production for up to 30 min in a dose-related manner, the effect being maximum at 0.1 mM concentration and decreasing at higher doses. The effect was blocked by ouabain (1 mM) as well as by the substitution of choline for Na+ in the incubation medium. Glutamate and aspartate exerted a less pronounced excitatory effect on CO2 production at higher concentration than kainate. This effect was also abolished by ouabain. Glutamate, added to the medium at a concentration at least 100-fold higher than kainate, partially reversed the increase in CO2 production induced by kainic acid. No similar effect was noticed for aspartate. The supposed glutamate antagonists glutamic acid diethylester (1 mM) and proline (5 mM) did not affect the excitatory action of kainic acid or exert an antagonistic effect towards glutamate. At higher concentration (10 mM) glutamic acid diethylester increased CO2 production, an effect that was, however, ouabain insensitive. Methyltetrahydrofolic acid (1 mM), a substance reported to compete for the kainate receptor, did not inhibit the effect of kainic acid or increase CO2 production.

Kainic acid neurotoxicity does not depend on intact retinal input in the goldfish optic tectum.

Kainic acid neurotoxicity has been studied in the optic tectum of the goldfish 4 weeks after eye enucleation. The effect of drug treatment has been tested with respect to both neurochemical and morphological parameters. The neurotransmitter-related enzymes, choline acetyltransferase, acetylcholinesterase and glutamate decarboxylase, show about 50% decrease in the deafferented tectum 6 days after kainic acid administration. Relevant morphological alterations of the tectal structure can also be noticed at the same stage. The neurotoxic effects of kainic acid in the deafferented optic tectum are therefore quite similar to the effects of previously noticed for the intact optic tectum of normal fish. Control experiments on the effect of optic nerve degeneration by itself on the levels of the neurotransmitter-related enzymes in the optic tectum, have shown no significant decrease in glutamate decarboxylase, a slight decrease in acetylcholinesterase and a more marked drop in choline acetyltransferase. The findings are discussed with reference to some of the hypotheses advanced in order to explain kainic acid neurotoxicity. It is proposed that the neurotoxic effect of kainic acid after removal of specific excitatory afferents, may vary in different nervous centers depending on differences of the remaining extrinsic connections and of the intrinsic neural circuits.