Agustín González

Catecholamine systems in the brain of vertebrates: new perspectives through a comparative approach.

A comparative analysis of catecholaminergic systems in the brain and spinal cord of vertebrates forces to reconsider several aspects of the organization of catecholamine systems. Evidence has been provided for the existence of extensive, putatively catecholaminergic cell groups in the spinal cord, the pretectum, the habenular region, and cortical and subcortical telencephalic areas. Moreover, putatively dopamine- and noradrenaline-accumulating cells have been demonstrated in the hypothalamic periventricular organ of almost every non-mammalian vertebrate studied. In contrast with the classical idea that the evolution of catecholamine systems is marked by an increase in complexity going from anamniotes to amniotes, it is now evident that the brains of anamniotes contain catecholaminergic cell groups, of which the counterparts in amniotes have lost the capacity to produce catecholamines. Moreover, a segmental approach in studying the organization of catecholaminergic systems is advocated. Such an approach has recently led to the conclusion that the chemoarchitecture and connections of the basal ganglia of anamniote and amniote tetrapods are largely comparable. This review has also brought together data about the distribution of receptors and catecholaminergic fibers as well as data about developmental aspects. From these data it has become clear that there is a good match between catecholaminergic fibers and receptors, but, at many places, volume transmission seems to play an important role. Finally, although the available data are still limited, striking differences are observed in the spatiotemporal sequence of appearance of catecholaminergic cell groups, in particular those in the retina and olfactory bulb.

Evolution of the basal ganglia in tetrapods: a new perspective based on recent studies in amphibians

It has been postulated frequently that the fundamental organization of the basal ganglia (BG) in vertebrates arose with the appearance of amniotes during evolution. An alternative hypothesis, however, is that such a condition was already present in early anamniotic tetrapods and, therefore, characterizes the acquisition of the tetrapod phenotype rather than the anamniotic-amniotic transition. Re-examination of the BG organization in tetrapods in the light of recent findings in amphibians strongly supports the notion that elementary BG structures were present in the brain of ancestral tetrapods and that they were organized according to a general plan shared today by all extant tetrapods.

Basal ganglia organization in amphibians: Chemoarchitecture

Recent studies dealing with the investigation of the afferent and efferent connections of the basal ganglia of amphibians have revealed many similarities with basal ganglia structures of amniotes. In a further step, the chemoarchitecture of basal ganglia of the frog Rana perezi has been investigated. For use as main markers of amphibian basal ganglia structures, antibodies against tyrosine hydroxylase, substance P, and enkephalin were selected. Moreover, the distributions of nitric oxide synthase (nicotinamide adenine dinucleotide phosphate‐diaphorase histochemistry), calretinin, dopamine‐β‐hydroxylase, choline acetyltransferase, mesotocin, vasotocin, somatostatin, neuropeptide Y, neuropeptide FF, and serotonin were studied to corroborate a comparison with both basal ganglia and amygdaloid structures of amniotes. On the basis of connections and chemoarchitecture, a striatum proper, nucleus accumbens, dorsal and ventral pallidum, bed nucleus of the stria terminalis, and amygdaloid complex have been identified. Accordingly, a new terminology is proposed that is in line with our current understanding of basal ganglia organization in amphibians. J. Comp. Neurol. 392:285–312, 1998.

Distribution of choline acetyltransferase immunoreactivity in the brain of anuran (Rana perezi, Xenopus laevis) and urodele (Pleurodeles waltl) amphibians.

Because our knowledge of cholinergic systems in the brains of amphibians is limited, the present study aimed to provide detailed information on the distribution of cholinergic cell bodies and fibers as revealed by immunohistochemistry with antibodies directed against the enzyme choline acetyltransferase (ChAT). To determine general and derived features of the cholinergic systems within the class of Amphibia, both anuran (Rana perezi, Xenopus laevis) and urodele (Pleurodeles waltl) amphibians were studied. Distinct groups of ChAT‐immunoreactive cell bodies were observed in the basal telencephalon, hypothalamus, habenula, isthmic nucleus, isthmic reticular formation, cranial nerve motor nuclei, and spinal cord. Prominent plexuses of cholinergic fibers were found in the olfactory bulb, pallium, basal telencephalon, ventral thalamus, tectum, and nucleus interpeduncularis. Comparison of these results with those obtained in other vertebrates, including a segmental approach to correlate cell populations, reveals that the cholinergic systems in amphibians share many features with amniotes. Thus, cholinergic pedunculopontine and laterodorsal tegmental nuclei could be identified in the amphibian brain. The finding of weakly immunoreactive cells in the striatum of Rana, which is in contrast with the condition found in Xenopus, Pleurodeles, and other anamniotes studied so far, has revived the notion that basal ganglia organization is more preserved during evolution than previously thought. J. Comp. Neurol. 382:499‐534, 1997.

Basal ganglia organization in amphibians: Afferent connections to the striatum and the nucleus accumbens

As part of a research program to determine if the organization of basal ganglia (BG) of amphibians is homologous to that of amniotes, the afferent connections of the BG in the anurans Xenopus laevis and Rana perezi and the urodele Pleurodeles waltl were investigated with sensitive tract‐tracing techniques. Hodological evidence is presented that supports a division of the amphibian BG into a nucleus accumbens and a striatum. Both structures have inputs in common from the olfactory bulb, medial pallium, striatopallial transition area, preoptic area, ventral thalamus, ventral hypothalamic nucleus, posterior tubercle, several mesencephalic and rhombencephalic reticular nuclei, locus coeruleus, raphe, and the nucleus of the solitary tract. Several nuclei that project to both subdivisions of the BG, however, show a clear preference for either the striatum (lateral amygdala, parabrachial nucleus) or the nucleus accumbens (medial amygdala, ventral midbrain tegmentum). In addition, the anterior entopeduncular nucleus, central thalamic nucleus, anterior and posteroventral divisions of the lateral thalamic nucleus, and torus semicircularis project exclusively to the striatum, whereas the anterior thalamic nucleus, anteroventral, and anterodorsal tegmental nuclei provide inputs solely to the nucleus accumbens. Apart from this subdivision of the basal forebrain, the results of the present study have revealed more elaborate patterns of afferent projections to the BG of amphibians than previously thought. Moreover, regional differences within the striatum and the nucleus accumbens were demonstrated, suggesting the existence of functional subdivisions. The present study has revealed that the organization of the afferent connections to the BG in amphibians is basically similar to that of amniotes. According to their afferent connections, the striatum and the nucleus accumbens of amphibians may play a key role in processing olfactory, visual, auditory, lateral line, and visceral information. However, contrary to the situation in amniotes, only a minor involvement of pallial structures on the BG functions is present in amphibians. J. Comp. Neurol. 378:16–49, 1997.

Distribution of choline acetyltransferase immunoreactivity in the brain of an elasmobranch, the lesser spotted dogfish (Scyliorhinus canicula).

Although the distribution of cholinergic cells is remarkably similar across the vertebrate species, no data are available on more primitive species, such as cartilaginous fishes. To extend the evolutionary analysis of the cholinergic systems, we studied the distribution of cholinergic neurons in the brain and rostral spinal cord of Scyliorhinus canicula by immunocytochemistry using an antibody against the enzyme choline acetyltransferase (ChAT). Western blot analysis of brain extracts of dogfish, sturgeon, trout, and rat showed that this antibody recognized similar bands in the four species. Putative cholinergic neurons were observed in most brain regions, including the telencephalon, diencephalon, cerebellum, and brainstem. In the retrobulbar region and superficial dorsal pallium of the telencephalon, numerous small pallial cells were ChAT‐like immunoreactive. In addition, tufted cells of the olfactory bulb and some cells in the lateral pallium showed faint immunoreactivity. In the preoptic‐hypothalamic region, ChAT‐immunoreactive (ChAT‐ir) cells were found in the preoptic nucleus, the vascular organ of the terminal lamina, and a small population in the caudal tuber. In the epithalamus, the pineal photoreceptors were intensely positive. Many cells of the habenula were faintly ChAT‐ir, but the neuropil of the interpeduncular nucleus showed intense ChAT immunoreactivity. In the pretectal region, ChAT‐ir cells were observed only in the superficial pretectal nucleus. In the brainstem, the somatomotor and branchiomotor nuclei, the octavolateral efferent nucleus, and a cell group just rostral to the Edinger‐Westphal (EW) nucleus contained ChAT‐ir neurons. In addition, the trigeminal mesencephalic nucleus, the nucleus G of the isthmus, some locus coeruleus cells, and some cell populations of the vestibular nuclei and of the electroreceptive nucleus of the octavolateral region exhibited ChAT immunoreactivity. In the reticular areas of the brainstem, the nucleus of the medial longitudinal fascicle, many reticular neurons of the rhombencephalon, and cells of the nucleus of the lateral funiculus were immunoreactive to this antibody. In the cerebellum, Golgi cells of the granule cell layer and some cells of the cerebellar nucleus were also ChAT‐ir. In the rostral spinal cord, ChAT immunoreactivity was observed in cells of the motor column, the dorsal horn, the marginal nucleus (a putative stretch‐receptor organ), and in interstitial cells of the ventral funiculus. These results demonstrate for the first time that cholinergic neurons are distributed widely in the central nervous system of elasmobranchs and that their cholinergic systems have evolved several characteristics that are unique to this group. J. Comp. Neurol. 420:139–170, 2000.

Distribution of choline acetyltransferase-immunoreactive structures in the lamprey brain

The distribution of cholinergic neurons and fibers was studied immunohistochemically in the brain of two species of lampreys (Petromyzon marinus and Lampetra fluviatilis), by using an antiserum against choline acetyltransferase (ChAT). The results obtained in both species were similar, but there appeared some interspecies differences. In the forebrain, cholinergic cells were present in the striatum, preoptic region, paraventricular nucleus, pineal and parapineal organs, habenula, and pretectum. The cranial nerve motoneurons (III, IV, V, VI, VII, IX, and X), the first and second spino‐occipital nerves (so), and the ventral horn of the spinal cord showed a strong ChAT immunoreactivity. Additional cholinergic neurons were observed: the mesencephalic M5 nucleus of Schober, two different cell populations in the isthmic region, the efferent component of the eighth nerve, putative preganglionic parasympathetic cells, cells in the solitary tract nucleus, and the rhombencephalic reticular formation. Cholinergic fibers were widely distributed in the brain. Comparison with previous studies in other vertebrates suggests that major cholinergic pathways, like tectal innervation from the isthmic region, are also present in lampreys. Of particular interest was the prominent projection to the neurohypophysis from cholinergic neurons in the preoptic region and paraventricular nucleus. Present data were analyzed within the segmental paradigm, as was previously done in other vertebrates. Our results reveal that the organization of many cholinergic systems in the lamprey as, for example, in the striatal, preoptic, and isthmic regions, comprises features of the anamniote brain that remain common to all living amniotes studied so far, thus being conservative to a surprisingly high degree. Therefore, the distribution of ChAT‐immunoreactive structures in the lamprey brain is, in general, comparable to that previously described in other vertebrate species. J. Comp. Neurol. 431:105–126, 2001.

Evolution of the basal ganglia: new perspectives through a comparative approach.

The basal ganglia (BG) have received much attention during the last 3 decades mainly because of their clinical relevance. Our understanding of their structure, organisation and function in terms of chemoarchitecture, compartmentalisation, connections and receptor localisation has increased equally. Most of the research has been focused on the mammalian BG, but a considerable number of studies have been carried out in nonmammalian vertebrates, in particular reptiles and birds. The BG of the latter 2 classes of vertebrates, which together with mammals constitute the amniotic vertebrates, have been thoroughly studied by means of tract‐tracing and immunohistochemical techniques. The terminology used for amniotic BG structures has frequently been adopted to indicate putative corresponding structures in the brain of anamniotes, i.e. amphibians and fishes, but data for such a comparison were, until recently, almost totally lacking. It has been proposed several times that the occurrence of well developed BG structures probably constitutes a landmark in the anamniote‐amniote transition. However, our recent studies of connections, chemoarchitecture and development of the basal forebrain of amphibians have revealed that tetrapod vertebrates share a common pattern of BG organisation. This pattern includes the existence of dorsal and ventral striatopallidal systems, reciprocal connections between the striatopallidal complex and the diencephalic and mesencephalic basal plate (striatonigral and nigrostriatal projections), and descending pathways from the striatopallidal system to the midbrain tectum and reticular formation. The connectional similarities are paralleled by similarities in the distribution of chemical markers of striatal and pallidal structures such as dopamine, substance P and enkephalin, as well as by similarities in development and expression of homeobox genes. On the other hand, a major evolutionary trend is the progressive involvement of the cortex in the processing of the thalamic sensory information relayed to the BG of tetrapods. By using the comparative approach, new insights have been gained with respect to certain features of the BG of vertebrates in general, such as the segmental organisation of the midbrain dopaminergic cell groups, the occurrence of large numbers of dopaminergic cell bodies within the telencephalon itself and the variability in, among others, connectivity and chemoarchitecture. However, the intriguing question whether the basal forebrain organisation of nontetrapods differs essentially from that observed in tetrapods still needs to be answered.

LIM-homeodomain genes as developmental and adult genetic markers of Xenopus forebrain functional subdivisions

We have investigated the expression patterns of five LIM‐homeodomain (LIM‐hd) genes, x‐Lhx1, x‐Lhx2, x‐Lhx5, x‐Lhx7, and x‐Lhx9 in the forebrain of the frog Xenopus laevis during larval development and in the adult. The results were analyzed in terms of neuromeric organization of the amphibian brain and of combinatorial LIM‐hd code and showed that LIM‐hd developmental transcription factors are particularly powerful to highlight the coherence of several groups or nuclei, to delineate subdivisions, and/or to clarify structures that are still a matter of debate. Among other findings, we bring substantial evidence for the following: (1) a dual origin of olfactory bulb neurons, based on x‐Lhx5 expression; (2) the existence of a ventral pallium in frog, based on x‐Lhx9 expression; (3) a multiple (pallial and subpallial) origin for the nuclei of the amygdaloid complex, based on distinct combinations of the five studied genes; (4) a clear homology between the Xenopus medial pallium and the mammalian hippocampus, based on x‐Lhx2 pattern; and (5) a confirmed prosomeric organization of the diencephalon, based on alternating x‐Lhx1/5 and x‐Lhx2/9 expressions. In addition, the important expression levels for LIM‐hd factors found throughout development and in the adult brain suggest a role for these genes in development and maintenance of neuronal specification and phenotype, as for example in the case of x‐Lhx7 and cholinergic neurons. Moreover, following LIM‐hd patterns throughout development points out to some of the migrations and morphogenetic movements, which give rise to the adult structures. Finally, the detailed description of the LIM‐hd code in the developing and adult Xenopus forebrain provides interesting cues for the possible mechanisms of evolution of the vertebrate forebrain. J. Comp. Neurol. 472:52–72, 2004.

Distribution of tyrosine hydroxylase and dopamine immunoreactivities in the brain of the South African clawed frog Xenopus laevis

SummaryThe distribution of dopamine (DA) and the biosynthetic enzyme tyrosine hydroxylase (TH) has been studied immunohistochemically in the brain of the adult South African clawed frog, Xenopus laevis. The goals of the present study are, firstly, to provide detailed information on the DA system of the brain of a species which is commonly used in laboratories as an experimental model and, secondly, to enhance our insight into primitive and derived characters of this catecholaminergic system in amphibians. Dopamine-immunoreactive cell bodies are present in the olfactory bulb, the preoptic area, the suprachiasmatic nucleus, the nucleus of the periventricular organ and its accompanying cells, the nucleus of the posterior tubercle, the posterior thalamic nucleus, the midbrain tegmentum, around the solitary tract, in the ependymal layer along the midline of the caudal rhombencephalon, and along the central canal of the spinal cord. In contrast to the DA antiserum, the TH antiserum fails to stain the liquor-contacting cells in the periventricular organ. On the contrary, the latter antiserum reveals additional immunoreactive cell bodies in the olfactory bulb, the isthmic region and the caudal brainstem. Both antisera yield an almost identical distribution of fibers. Distinct fiber plexuses are observed in the olfactory bulb, the basal forebrain, the hypothalamus and the intermediate lobe of the hypophysis. Features that Xenopus shares with other anurans are the larger number of DAi cells, which are generally smaller in size than those observed in urodeles, and the lack of DAi fibers in pallial structures. On the other hand, the paired midbrain DA cell group and the innervation of the tectum of Xenopus resemble those found in the newt rather than those in frogs. Despite the existence of these species differences, the brain of Xenopus offers an excellent model for studying general aspects of neurotransmitter interactions and the development of catecholamine systems in this class of vertebrates.