Skirmantas Janušonis

Early serotonergic projections to Cajal-Retzius cells: relevance for cortical development.

Although the serotonergic system plays an important role in various neurological disorders, the role of early serotonergic projections to the developing cerebral cortex is not well understood. Because serotonergic fibers enter the marginal zone (MZ) before birth, it has been suggested that they may influence cortical development through synaptic contacts with Cajal-Retzius (CR) cells. We used immunohistochemistry combined with confocal and electron microscopy to show that the earliest serotonergic projections to the MZ form synaptic contacts with the somata and proximal dendrites of CR cells as early as embryonic day 17. To elucidate the functional significance of these early serotonergic contacts with CR cells, we perturbed their normal development by injecting pregnant mice with 5-methoxytryptamine. Lower reelin levels were detected in the brains of newborn pups from the exposed animals. Because reelin plays an important role in the cortical laminar and columnar organization during development, we killed some pups from the same litters on postnatal day 7 and analyzed their presubicular cortex. We found that the supragranular layers of the presubicular cortex (which normally display a visible columnar deployment of neurons) were altered in the treated animals. Our results suggest a mechanism of how serotonergic abnormalities during cortical development may disturb the normal cortical organization; and, therefore, may be relevant for understanding neurological disorders in which abnormalities of the serotonergic system are accompanied by cortical pathology (such as autism).

Retinal afferents to the dorsal raphe nucleus in rats and Mongolian gerbils

A direct pathway from the retina to the dorsal raphe nucleus (DRN) has been demonstrated in both albino rats and Mongolian gerbils. Following intraocular injection of cholera toxin subunit B (CTB), a diffuse stream of CTB‐positive, fine‐caliber optic axons emerged from the optic tract at the level of the pretectum/anterior mesencephalon. In gerbils, CTB‐positive axons descended ventromedially into the periaqueductal gray, moving caudally and arborizing extensively throughout the DRN. In rats, the retinal‐DRN projection comprised fewer, but larger caliber, axons, which arborized in a relatively restricted region of the lateral and ventral DRN. Following injection of CTB into the lateral DRN, retrogradely labeled ganglion cells (GCs) were observed in whole‐mount retinas of both species. In gerbils, CTB‐positive GCs were distributed over the entire retina, and a nearest‐neighbor analysis of CTB‐positive GCs showed significant regularity (nonrandomness) in their distribution. The overall distribution of gerbil GC soma diameters ranged from 8 to 22 μm and was skewed slightly towards the larger soma diameters. Based on an adaptive mixtures model statistical analysis, two Gaussian distributions appeared to comprise the total GC distribution, with mean soma diameters of 13 (SEM ±1.7) μm, and 17 (SEM ±1.5) μm, respectively. In rats, many fewer CTB‐positive GCs were labeled following CTB injections into the lateral DRN, and nearly all occurred in the inferior retina. The total distribution of rat GC soma diameters was similar to that in gerbils and also was skewed towards the larger soma diameters. Major differences observed in the extent and configuration of the retinal‐DRN pathway may be related to the diurnal/crepuscular vs. nocturnal habits of these two species. J. Comp. Neurol. 414:469–484, 1999.

Retinal projection to the dorsal raphe nucleus in the Chilean degus (Octodon degus)

A substantial projection from the retina to the dorsal raphe nucleus (DRN) has been demonstrated in the Chilean degus, a diurnal/crepuscular hystricomorph rodent. Following intraocular injection of cholera toxin subunit B (CTB), immunocytochemically labeled CTB-positive axons and terminals were observed in all major retinorecipient nuclei as well as in the DRN and periaqueductal gray (PAG) of the mesencephalon. Two streams of optic axons to the DRN were observed: one descending from the optic tract at the level of the pretectum and anterior superior colliculus, the other emerging as a small fascicle at the anterior pole of the inferior colliculus and descending bilaterally through the PAG. Contralateral retinal afferents in the DRN appeared to terminate primarily in the dorsomedial and lateral subdivisions of the DRN, and a less extensive ipsilateral component also was observed. Axonal arborizations were characterized by short branches and multiple varicosities, both in the DRN and in the PAG. The extent and density of DRN retinal afferents were not as extensive as previously observed in Mongolian gerbils using identical techniques, but the retinal-DRN projection is considerably larger in degus than in rats. The functional significance of the retinal-DRN pathway remains to be determined, although a variety of evidence indicates that light may directly affect the activity of neurons and serotonin levels in the DRN.

Religion priming differentially increases prosocial behavior among variants of the dopamine D4 receptor (DRD4) gene

Building on gene-environment interaction (G × E) research, this study examines how the dopamine D4 receptor (DRD4) gene interacts with a situational prime of religion to influence prosocial behavior. Some DRD4 variants tend to be more susceptible to environmental influences, whereas other variants are less susceptible. Thus, certain life environments may be associated with acts of prosociality for some DRD4 variants but not others. Given that religion can act as an environmental influence that increases prosocial behavior, environmental input in the form of religion priming may have G × E effects. Results showed that participants with DRD4 susceptibility variants were more prosocial when implicitly primed with religion than not primed with religion, whereas participants without DRD4 susceptibility variants were not impacted by priming. This research has implications for understanding why different people may behave prosocially for different reasons and also integrates G × E research with experimental psychology.

Ontogeny of brain and blood serotonin levels in 5‐HT1A receptor knockout mice: potential relevance to the neurobiology of autism

The most consistent neurochemical finding in autism has been elevated group mean levels of blood platelet 5‐hydroxytryptamine (5‐HT, serotonin). The origin and significance of this platelet hyperserotonemia remain poorly understood. The 5‐HT1A receptor plays important roles in the developing brain and is also expressed in the gut, the main source of platelet 5‐HT. Post‐natal tissue levels of 5‐HT, 5‐hydroxyindoleacetic acid (5‐HIAA) and tryptophan were examined in the brain, duodenum and blood of 5‐HT1A receptor‐knockout and wild‐type mice. At 3 days after birth, the knockout mice had lower mean brain 5‐HT levels and normal mean platelet 5‐HT levels. Also, at 3 days after birth, the mean tryptophan levels in the brain, duodenum and blood of the knockout mice were around 30% lower than those of the wild‐type mice. By 2 weeks after birth, the mean brain 5‐HT levels of the knockout mice normalized, but their mean platelet 5‐HT levels became 24% higher than normal. The possible causes of these dynamic shifts were explored by examining correlations between central and peripheral levels of 5‐HT, 5‐HIAA and tryptophan. The results are discussed in relation to the possible role of 5‐HT in the ontogeny of autism.

Origin of the blood hyperserotonemia of autism

BackgroundResearch in the last fifty years has shown that many autistic individuals have elevated serotonin (5-hydroxytryptamine, 5-HT) levels in blood platelets. This phenomenon, known as the platelet hyperserotonemia of autism, is considered to be one of the most well-replicated findings in biological psychiatry. Its replicability suggests that many of the genes involved in autism affect a small number of biological networks. These networks may also play a role in the early development of the autistic brain.ResultsWe developed an equation that allows calculation of platelet 5-HT concentration as a function of measurable biological parameters. It also provides information about the sensitivity of platelet 5-HT levels to each of the parameters and their interactions.ConclusionThe model yields platelet 5-HT concentrations that are consistent with values reported in experimental studies. If the parameters are considered independent, the model predicts that platelet 5-HT levels should be sensitive to changes in the platelet 5-HT uptake rate constant, the proportion of free 5-HT cleared in the liver and lungs, the gut 5-HT production rate and its regulation, and the volume of the gut wall. Linear and non-linear interactions among these and other parameters are specified in the equation, which may facilitate the design and interpretation of experimental studies.

Comparing two small samples with an unstable, treatment-independent baseline.

Due to time and resource constraints, small samples (N=3-7 cases per group) are often used in neurobiological studies that employ multiple techniques. In a simulation study, five statistical tests were used to compare two small samples (treated and control) with an unstable, additive baseline. These five tests differed in the way that they used the baseline variable (B) to adjust or normalize the variable affected by the treatment (Y). We conclude that, if N=3 or 4, the independent t-test on Y-B tends to have the highest power; if N> or =7, ANCOVA on Y with B as the covariate tends to have the highest power; and both tests have comparably high power if N=5 or 6. The Wilcoxon rank-sum test (or, equivalently, the Mann-Whitney test) has precisely zero power if one group has 3 cases and the other has 3 or 4 cases. Some other problems of small-sample analysis are considered.

Distribution of the GABAA receptor complex β2/3 subunits in the brain of the frog Rana pipiens

This report describes the distribution of labeling of the monoclonal antibody bd-17 against the beta 2/3 subunits of the mammalian GABAA receptor complex throughout the brain of the frog Rana pipiens. The distribution matches quite closely those in homologous brain regions as previously described for this antibody in fishes, birds, and mammals, indicating that this antibody also labels beta 2/3 subunits of frog. A semiquantitative analysis of the distribution of labeling throughout the brain is based upon relative optical densities with respect to the structure showing maximal optical density in each brain, using standard illumination conditions. Comparison with distributions in birds and mammals suggests that these GABAA receptor complex subunits are strongly conserved in vertebrate evolution and play an important role in the visual, auditory, olfactory and motor systems.

Retinal ganglion cells projecting to the dorsal raphe and lateral geniculate complex in Mongolian gerbils.

Injections of rhodamine-B into the dorsal raphe nucleus (DRN) and Fluoro-Gold into the lateral geniculate nucleus (LGN) revealed double-labeled retinal ganglion cells (DL RGCs) projecting to both nuclei. The soma-size distribution of DL RGCs was compared with three other distributions: DRN-projecting RGCs, LGN-projecting RGCs, and a large sample of RGCs labeled via the optic nerve with DiI. DL RGC soma diameters fell primarily within the mid-to-upper size range of all three distributions. DL RGCs may provide information to both nuclei concerning comparable aspects of light and visual stimulation via collateralized axons.

BRAIN GROWTH TRAJECTORIES IN MOUSE STRAINS WITH CENTRAL AND PERIPHERAL SEROTONIN DIFFERENCES: RELEVANCE TO AUTISM MODELS

The genetic heterogeneity of autism spectrum disorders (ASDs) suggests that their underlying neurobiology involves dysfunction at the neural network level. Understanding these neural networks will require a major collaborative effort and will depend on validated and widely accepted animal models. Many mouse models have been proposed in autism research, but the assessment of their validity often has been limited to measuring social interactions. However, two other well-replicated findings have been reported in ASDs: transient brain overgrowth in early postnatal life and elevated 5-HT (serotonin) levels in blood platelets (platelet hyperserotonemia). We examined two inbred mouse strains (C57BL/6 and BALB/c) with respect to these phenomena. The BALB/c strain is less social and exhibits some other autistic-like behaviors. In addition, it has a lower 5-HT synthesis rate in the central nervous system due to a single-nucleotide polymorphism in the tryptophan hydroxylase 2 (Tph2) gene. The postnatal growth of brain mass was analyzed with mixed-effects models that included litter effects. The volume of the hippocampal complex and the thickness of the somatosensory cortex were measured in 3D-brain reconstructions from serial sections. The postnatal whole-blood 5-HT levels were assessed with high-performance liquid chromatography. With respect to the BALB/c strain, the C57BL/6 strain showed transient brain overgrowth and persistent blood hyperserotonemia. The hippocampal volume was permanently enlarged in the C57BL/6 strain, with no change in the adult brain mass. These results indicate that, in mice, autistic-like shifts in the brain and periphery may be associated with less autistic-like behaviors. Importantly, they suggest that consistency among behavioral, anatomical, and physiological measures may expedite the validation of new and previously proposed mouse models of autism, and that the construct validity of models should be demonstrated when these measures are inconsistent.