Altered development of dopaminergic cells in the retina of weaver mice.

Postnatal degeneration of dopaminergic (DA) cells is known to occur in mesencephalic nuclei of mutant weaver mice, whereas retinal DA content is reported to be unchanged in the adult animal. To determine whether morphological changes occur in the weaver retinal DA system, we compared weaver and control developing and adult retinas after tyrosine hydroxylase (TH) immunohistochemistry. The density and distribution of DA cells were analyzed using Dirichlet tessellation. Not only was no DA cell loss found in adult weaver retinas, but we even observed an increase in DA cells in weaver compared to control retinas between postnatal days 14 and 30. Furthermore, some unusual features were found during the latter period: atypical cells (representing a maximum of 12% of the whole DA cell population) were observed, and these differed from typical DA cells in terms of both location (slightly more external within the inner nuclear layer) and appearance (flat somata, round and clear nuclei, thick dendritic trunks emerging laterally and giving rise to horizontal processes). Some of the atypical cells were intermingled in a delicate network lying in a more outer focal plane than the main DA plexus. The expression of GIRK2, a G protein-related inward rectifying K(+) channel responsible for the weaver syndrome, was investigated. Although no GIRK2 labeling was demonstrated in DA cells, its possible involvement in the transient disturbances observed in the weaver DA retinal system is discussed.

An immunocytochemical study on the distribution of two G-protein-gated inward rectifier potassium channels (GIRK2 and GIRK4) in the adult rat brain.

G-protein-gated inward rectifier potassium channels mediate the synaptic actions of numerous neurotransmitters in the mammalian brain, and were recently shown to be candidates for genetic mutations leading to neuronal cell death. This report describes the localization of G-protein-gated inward rectifier potassium channel-2 and G-protein-gated inward rectifier potassium channel-4 proteins in the rat brain, as assessed by immunocytochemistry. G-protein-gated inward rectifier potassium channel-2 immunoreactivity was widely distributed throughout the brain, with the strongest staining seen in the hippocampus, septum, granule cell layer of the cerebellum, amygdala and substantia nigra pars compacta. In contrast, G-protein-gated inward rectifier potassium channel-4 immunoreactivity was restricted to some neuronal populations, such as Purkinje cells and neurons of the globus pallidus and the ventral pallidum. The presence of G-protein-gated inward rectifier potassium channel-2 immunoreactivity in substantia nigra pars compacta dopaminergic neurons was confirmed by showing its co-localization with tyrosine hydroxylase by double immunocytochemistry, and also by selectively lesioning dopaminergic neurons with the neurotoxin 6-hydroxydopamine. At the cellular level both proteins were localized in neuronal cell bodies and dendrites, but clear differences were seen in the degree of dendritic staining among neuronal groups. For some neuronal groups the staining of distal dendrites (notably dendritic spines) was strong, while for others the cell body and proximal dendrites were preferentially labelled. In addition, some of the results suggest that G-protein-gated inward rectifier potassium channel-2 protein could be localized in distal axonal terminal fields. A knowledge of the distribution of G-protein-gated inward rectifier potassium channel proteins in the brain could help to elucidate their physiological roles and to evaluate their potential involvement in neurodegenerative processes in animal models and human diseases.

Comparative expression of the inward rectifier K+ channel GIRK2 in the cerebellum of normal and weaver mutant mice.

The main target for degeneration associated with the weaver mutation is the cerebellum. Expression of the GIRK2 mRNA and protein was studied in cerebellum of 12- and 22-day-old normal and weaver mice. In 12-day-old mice, GIRK2 is expressed at highest levels in the external granule layer (EGL) and in lower levels in the newly forming internal granule layer (IGL). In the weaver cerebellum, a high hybridization signal and dark immunostaining was observed in the EGL due to the higher density of non-migrated cells. In 22-day-old weaver cerebella, there are only few remaining granule cells existing as scattered cells within the IGL and molecular layer. GIRK2 is expressed in these neurons but the majority of cells expressing GIRK2 in these cerebella are Purkinje cells that are also affected by the weaver mutation (position, shape) but have not died. Normal cerebellar granule neurons but not homozygous mutant neurons in primary cultures and cerebellar slices of 8-day-old mice displayed inward rectifier K+ currents. Taken together, these findings suggest that cell loss in the weaver cerebellum is not directly related to a differential content of GIRK2 in the affected neurons during development. The lethal effect of the weaver mutation in specific neurons is probably due to a combination of the abnormal function of the inward rectifier K+ channels and other factors specific to the vulnerable neurons.

An immunocytochemical study of a G-protein-gated inward rectifier K+ channel (GIRK2) in the weaver mouse mesencephalon.

It has been suggested that a mutation in a G-protein-gated inward rectifier K+ channel (GIRK2) is responsible for inducing cell death in the cerebellum of homozygous weaver (wv/wv) mutant mice. These mice also display a progressive, massive loss of mesencephalic dopaminergic neurones. Using an immunocytochemical method, we detected GIRK2-positive cell bodies and fibres in the substantia nigra pars compacta (SNC) and the ventral tegmental area (VTA) of control (+/+) mice. Cell counts of both GIRK2- and tyrosine hydroxylase (TH)-positive neurones demonstrated a marked loss of SNC cell bodies, especially in 12-month-old (12M) wv/wv mice. A considerable proportion of GIRK2-positive cell bodies were preserved, however. In addition, no loss of GIRK2-positive neurones was observed in the VTA of 12M wv/wv mice, despite of a significant reduction in TH-positive cell bodies. These results suggest that expression of the mutated channel is not a sufficient condition to induce cell death in the ventral mesencephalon of the wv/wv mice.

Effect of the weaver mutation on the expression of dopamine membrane transporter, tyrosine hydroxylase and vesicular monoamine transporter in dopaminergic neurons of the substantia nigra and the ventral tegmental area.

The adult homozygous weaver mutant mouse (wv/wv) is characterized by a loss of dopamine (DA) neurons in the nigrostriatal pathway. Quantitative in situ hybridization of three different dopaminergic markers: dopamine membrane transporter (DAT), tyrosine hydroxylase (TH), and vesicular monoamine transporter (VMAT2) was performed on individual dopaminergic cells of the substantia nigra pars compacta (SNC) and the ventral tegmental area (VTA) in 2-month-old wv/wv mice, in order to investigate the metabolic state of remaining dopaminergic cell bodies and gain further insight into modifications observed on dopaminergic nerve terminals in the striatum and the nucleus accumbens. Cellular expression of DAT mRNA in remaining dopaminergic cells of both the SNC and the VTA was decreased in the wv/wv mice compared to the wild-type mice (+/+). In contrast, the expression of TH and VMAT2 mRNA remained unchanged in the wv/wv mice. Furthermore, in 7-day-old wv/wv mice, before the onset of cell death in the midbrain. DAT mRNA levels were reduced in dopaminergic neurons in both the SNC and VTA. In these animals, the cellular expression of TH mRNA remained unchanged. These results taken together indicate that DAT expression is one of the first targets in the ventral mesencephalon of the wv mutation, inducing a specific decrease of DA uptake in the striatum and the nucleus accumbens. The alteration of the DA membrane transporter could play a role in the progression of DA neuronal death in the wv mice.

Transient uptake and storage of serotonin in developing thalamic neurons.

Serotonin (5-HT) has been shown to affect the development and patterning of the mouse barrelfield. We show that the dense transient 5-HT innervation of the somatosensory, visual, and auditory cortices originates in the thalamus rather than in the raphe: 5-HT is detected in thalamocortical fibers and most 5-HT cortical labeling disappears after thalamic lesions. Thalamic neurons do not synthesize 5-HT but take up exogenous 5-HT through 5-HT high affinity uptake sites located on thalamocortical axons and terminals. 3H-5-HT injected into the cortex is retrogradely transported to thalamic neurons. In situ hybridization shows a transient expression of the genes encoding the serotonin transporter and the vesicular monoamine transporter in thalamic sensory neurons. In these glutamatergic neurons, internalized 5-HT might thus be stored and used as a "borrowed transmitter" for extraneuronal signaling or could exert an intraneuronal control on thalamic maturation.

Association of autosomal loci with the grooming activity in mice observed in open-field.

We studied the reactions of mice when placed in an open-field environment and counted the grooming score in response to novelty. We used animals from Mendelian F2s and backcrosses, obtained from the parental strains ABP/Le and C57BL/6By, to test the hypothesis that the differences in this behavior were due to genetic variation at loci associated with visible recessive markers. Furthermore, the analysis of the segregating populations by means of non parametric Collins' method was used to test the one segregating unit hypothesis. We provide evidence for genes involved in the variation of grooming activity and situated close to the locus se on chromosome 9. Although the one-locus hypothesis was ruled out in the general analysis, a possible major gene effect in some crosses suggests a maternal environment effect.