Uln, Univ., Diss., 2000.

Idiopathic Parkinson??s disease (PD) and the atypical parkinsonian syndromes progressive supranuclear palsy (PSP) and multiple system atrophy (MSA) have substantial overlap in clinical features, with parkinsonian and cerebellar phenotypes identified. Pathologically, reactive glial changes occur in overlapping pathways in these disorders. Recent findings that glia concentrate proteins associated with genetic forms of PD, suggest a more significant role for these cells in the pathogenesis of these disorders. This study is the first to comparatively assess glial abnormalities in PD, PSP and MSA. Cases were clinically and pathologically characterised to establish correlates to prominent glial changes. The three main types of glia (astrocytes, oligodendroglia and microglia) were characterised by morphological features and protein expression (using immunohistochemical techniques) for correlations with disease indices. Tissue features measured included subregional volumes, nigral cell loss, characteristic disease inclusions, and the densities of glia. Using these techniques, the parkin co-regulated gene protein was found to be a novel constituent protein in protoplasmic astrocytes, facilitating the assessment of glial subtypes. The data show that distinct glial abnormalities associate with each parkinsonian syndrome. The most marked differences were observed in the astrocytic reaction in each disorder. In PD, protoplasmic astrocytes accumulated non-fibrillar -synuclein and degenerated over time, relating to a loss of levodopa responsiveness. In PSP, there was a marked protoplasmic astrogliosis (with these reactive glia strongly expressing parkin) that related to PSP-prominent clinical symptoms. In MSA, there was a marked fibrous astrogliosis and a loss of protoplasmic astrocytes in association with early changes in constituent myelin and oligodendroglial proteins. In contrast, similar microglial activation was observed across all disorders, with phagocytes concentrating in the substantia nigra, while non-phagocytic reactive microglia expressed parkin and associated with inclusion formation in each disorder. Overall, the glial reactions were considered to be either contributing to or ameliorating (neuroprotective) the neurodegenerative processes, and the timing of these reactions assessed with respect to indices of disease progression. The novel findings of this thesis show that glial abnormalities are prominent but distinct, and occur early in these parkinsonian syndromes. Suggestions on how these findings may translate into future therapeutic targets are given.

Idiopathic Parkinson??s disease (PD) and the atypical parkinsonian syndromes progressive supranuclear palsy (PSP) and multiple system atrophy (MSA) have substantial overlap in clinical features, with parkinsonian and cerebellar phenotypes identified. Pathologically, reactive glial changes occur in overlapping pathways in these disorders. Recent findings that glia concentrate proteins associated with genetic forms of PD, suggest a more significant role for these cells in the pathogenesis of these disorders. This study is the first to comparatively assess glial abnormalities in PD, PSP and MSA. Cases were clinically and pathologically characterised to establish correlates to prominent glial changes. The three main types of glia (astrocytes, oligodendroglia and microglia) were characterised by morphological features and protein expression (using immunohistochemical techniques) for correlations with disease indices. Tissue features measured included subregional volumes, nigral cell loss, characteristic disease inclusions, and the densities of glia. Using these techniques, the parkin co-regulated gene protein was found to be a novel constituent protein in protoplasmic astrocytes, facilitating the assessment of glial subtypes. The data show that distinct glial abnormalities associate with each parkinsonian syndrome. The most marked differences were observed in the astrocytic reaction in each disorder. In PD, protoplasmic astrocytes accumulated non-fibrillar -synuclein and degenerated over time, relating to a loss of levodopa responsiveness. In PSP, there was a marked protoplasmic astrogliosis (with these reactive glia strongly expressing parkin) that related to PSP-prominent clinical symptoms. In MSA, there was a marked fibrous astrogliosis and a loss of protoplasmic astrocytes in association with early changes in constituent myelin and oligodendroglial proteins. In contrast, similar microglial activation was observed across all disorders, with phagocytes concentrating in the substantia nigra, while non-phagocytic reactive microglia expressed parkin and associated with inclusion formation in each disorder. Overall, the glial reactions were considered to be either contributing to or ameliorating (neuroprotective) the neurodegenerative processes, and the timing of these reactions assessed with respect to indices of disease progression. The novel findings of this thesis show that glial abnormalities are prominent but distinct, and occur early in these parkinsonian syndromes. Suggestions on how these findings may translate into future therapeutic targets are given.

No description available.

Morphine is one of the most commonly prescribed medications for the relief of prolonged pain. Both basic science and clinical studies indicate that females require 2-3 times more morphine than males to achieve the same analgesic effect. To date, the mechanisms underlying sex differences in opiate responsiveness are unknown. Recent studies suggest that glial cells are potent modulators of morphine-based analgesia, and in particular, decrease the analgesic effect of opiates. Therefore, we tested the hypothesis that the sexually dimorphic effects of morphine were due to sex differences in glial cell activity. Our studies focused on the midbrain periaqueductal gray (PAG) as this region of the brain is critical for the analgesic effects of morphine. Adult male and female Sprague Dawley rats (250g- 400g) were procured from Charles River Laboratories, and were allowed 7 days to acclimate to the new facility. On the day of the experiment, animals received a subcutaneous injection of morphine (5mg/kg) or were handled in a similar manner. Thirty or 60 minutes after injections or handling, animals were perfused with a 4% paraformaldehyde and 2.5% acrolein tissue fixative solution. Brains were removed and stored in 20% sucrose until ready for sectioning. Brains were sectioned at 25mm using a freezing microtome, and immunohistochemical localization of markers for astrocyte glial cell activity was performed. Antibodies to glial fibrillary acidic protein (GFAP) were used to label activated astrocytes. If our hypothesis is correct, then females will have significantly greater density of the astrocyte cell activity marker GFAP as compared with males. Sex differences in PAG glial cell activity may provide the biological bases for the sexually dimorphic effect of morphine. This research may lead to better treatment for females experiencing prolonged chronic or neuropathic pain.

Morphine is one of the most commonly prescribed medications for the relief of prolonged pain. Both basic science and clinical studies indicate that females require 2-3 times more morphine than males to achieve the same analgesic effect. To date, the mechanisms underlying sex differences in opiate responsiveness are unknown. Recent studies suggest that glial cells are potent modulators of morphine-based analgesia, and in particular, decrease the analgesic effect of opiates. Therefore, we tested the hypothesis that the sexually dimorphic effects of morphine were due to sex differences in glial cell activity. Our studies focused on the midbrain periaqueductal gray (PAG) as this region of the brain is critical for the analgesic effects of morphine. Adult male and female Sprague Dawley rats (250g- 400g) were procured from Charles River Laboratories, and were allowed 7 days to acclimate to the new facility. On the day of the experiment, animals received a subcutaneous injection of morphine (5mg/kg) or were handled in a similar manner. Thirty or 60 minutes after injections or handling, animals were perfused with a 4% paraformaldehyde and 2.5% acrolein tissue fixative solution. Brains were removed and stored in 20% sucrose until ready for sectioning. Brains were sectioned at 25mm using a freezing microtome, and immunohistochemical localization of markers for astrocyte glial cell activity was performed. Antibodies to glial fibrillary acidic protein (GFAP) were used to label activated astrocytes. If our hypothesis is correct, then females will have significantly greater density of the astrocyte cell activity marker GFAP as compared with males. Sex differences in PAG glial cell activity may provide the biological bases for the sexually dimorphic effect of morphine. This research may lead to better treatment for females experiencing prolonged chronic or neuropathic pain.

This study investigated the position, ultrastructure and life history of glia in the midline of the Drosophila melanogaster Central Nervous System (CNS) by using enhancer traps (AA142, X81, argoswll, pointed1277) and reporter constructs (EEl, slilacZ 1.0, slilacZ 4.5) as glial-specific markers. Previous work has established that glia are necessary for proper formation and morphogenesis of longitudinal and commissural axon tracts, and axon ensheathment (Jacobs, 1993; Klambt et al., 1991; Jacobs and Goodman, 1989). By the end of embryogenesis there are three midline glial (MG) cells remaining in each segment (Sonnenfeld and jacobs, in press) which this study verifies. In the third instar larval MG cells proliferate to 24 cells per segment as followed with the E. coli lacZ expressing pointed1277 strain. These E. coli lacZ expressing pointed1277 MG cells begin dividing 57 hours after hatching as seen with 5 -bromodeoxyuridine and hydroxyurea treatment. Some MG genes cease midline expression before MG proliferation (seen with EEl, X81), others (seen with AA142) continue to be expressed until the beginning of MG proliferation. Only the argoswll, slilacZ 1.0, slilacZ 4.5, and pointed1277 expression strains continue E. coli lacZ expression to the end of the larval stages. In the first larval stage a few perineuropilar glia begin to express the E. coli lacZ gene and increase to 400 cells per CNS in the third ins tar as seen in the pointed1277 marker strain. pointed1277 EM micrographs show that E. coli lacZ labeled cells have a glial-like ultrastructure. There - was no co-localization of the E. coli lacZ expression in pointed12 77 and an anti-RK2 (repo) antibody in third instar larvae. In pointed1277 pupae the MG cell E. coli lacZ expression stops after 48- 72 hours and the prerineuropilar signal stops after 24 hours. In newly hatched pointed1277 adults perineuropilar E. coli lacZ expression is present with a cluster of 12 cells in the center of the neuropil. To summarize, after embryogenesis, in the pointed1277 marker strain, the MG cells begin dividing after 57 hours and the E. coli lacZ gene expression ends after the second day of the pupal stage. In the first instar, perineuropilar glia begin to label for the E. coli lacZ product and this expression ends by one day into the pupal stage, with re-appearance in the adult CNS. / Thesis / Master of Science (MSc)

In Drosophila melanogaster, the genes of the spitz group and of the DER signaling pathway function together to communicate localized developmental signals to the cells of many tissues. The embryonic midline glia (MG), a mesectodermal lineage essential to proper morphogenesis of the axon tracts of the ventral nerve cord (VNC), depend on spitz group signaling for survival and differentiation. Loss of function of any of the spitz group genes or of DER results in a decrease in the number of MG and subsequent defects in the formation of the axon tracts. These defects include a medial collapse of the longitudinal axons and fusion of the posterior and anterior commissures. Ectopic expression of Rhomboid, a putative seven pass transmembrane protein which is a member of the spitz group, generates supernumerary glia. Directed expression of DER^AB87T, an activated form of the Drosophila EGF receptor, sSpi, a diffusible ligand, or Ras^v12, a constitutively activated variant of a monomeric G-protein, have the same effect on MG number. It is proposed that the spitz group/DER signaling pathway act to promote survival of MG precursor cells. In addition, expression of Draf and pntP1 are found to increase the number of surviving supernumerary MG, however these signaling molecules are determined to be less effective at promoting the survival of the MG, based on their decreased ability to generate supernumerary MG. Furthermore, a subset of the supernumerary MG created in embryos in which Draf has been misexpressed appear to be incompletely differentiated or insulated from normal programmed cell death (PCD) by expression of this transgene. The effect of multiple cytoplasmic signaling pathways, activated by spitz group/DER signaling, on MG survival and differentiation is examined and discussed. Ectopic expression of rhomboid generates supernumerary MG. This process is suppressed by mutation of genes of the spitz group or of DER. Evidence that Rho functions upstream of Spi, Star and DER is presented. The implications of these data and their relevance to previously published models which propose the molecular actions of the spitz group/DER pathway genes are discussed. Finally, the role of spitz group/DER signaling in the activation of downstream target genes is explored. Overexpression of these genes results in increased expression of pointed, argos, and rhomboid. A model for spitz group function in signal amplification is proposed. / Thesis / Master of Science (MSc)

The visual system of the fruit fly is ideal for studying the association between neurons and glia. These interact during the morphogenesis of brain neuropiles and, in the adult, work together to maintain an ideal environment for neuronal function. In this thesis I characterise the pupal metamorphosis and adult structure of glia in the optic lobe’s lamina and medulla neuropiles. Photoreceptor axons from the fly’s compound eyes terminate at locations within these neuropiles that allow them intimate contact with glia. Some neuropile glia take up and inactivate the neurotransmitter histamine after its release at photoreceptor synapses. A shuttle pathway between the glia and photoreceptors then transports inactivated histamine back to photoreceptors for reuse. The gene CG12120 encodes the protein Tan, which liberates recycled histamine within the photoreceptor cytoplasm, it is then pumped into vesicles for re-release. Histamine, however, is not exclusive to the visual system. A system of glial barriers in the lamina, and around the brain, controls the movement of histamine between neuropiles as well as between the body and the brain. How histamine is reciprocally transported between photoreceptors and glia remains unknown despite attempts to uncover candidate transporters. Photoreceptor-specific capitate projections form as invaginations from neuropile glia into photoreceptor terminals. The transmitter needs of the photoreceptor appear to dictate the dynamic structure of capitate projections, which change in shape and number after perturbations that affect terminal location, synapse number or histamine release at the photoreceptor. This dynamism suggests that capitate projections play an important role, not only in recycling synaptic vesicles, but also in recycling histamine.

No description available.