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F D Miller

Publications and source records attributed to F D Miller.

At least 37 records · Page 2Linked to original sources
PubMed1999

Depolarization and neurotrophins converge on the phosphatidylinositol 3-kinase-Akt pathway to synergistically regulate neuronal survival.

In this report, we have examined the mechanisms whereby neurotrophins and neural activity coordinately regulate neuronal survival, focussing on sympathetic neurons, which require target-derived NGF and neural activity for survival during development. When sympathetic neurons were maintained in suboptimal concentrations of NGF, coincident depolarization with concentrations of KCl that on their own had no survival effect, synergistically enhanced survival. Biochemical analysis revealed that depolarization was sufficient to activate a Ras-phosphatidylinositol 3-kinase-Akt pathway (Ras-PI3-kinase-Akt), and function-blocking experiments using recombinant adenovirus indicated that this pathway was essential for approximately 50% of depolarization-mediated neuronal survival. At concentrations of NGF and KCl that promoted synergistic survival, these two stimuli converged to promote increased PI3-kinase-dependent Akt phosphorylation. This convergent PI3-kinase-Akt pathway was essential for synergistic survival. In contrast, inhibition of calcium/calmodulin-dependent protein kinase II revealed that, while this molecule was essential for depolarization-induced survival, it had no role in KCl- induced Akt phosphorylation, nor was it important for synergistic survival by NGF and KCl. Thus, NGF and depolarization together mediate survival of sympathetic neurons via intracellular convergence on a Ras-PI3-kinase-Akt pathway. This convergent regulation of Akt may provide a general mechanism for coordinating the effects of growth factors and neural activity on neuronal survival throughout the nervous system.

Animals↗
PubMed1999

Functionally antagonistic interactions between the TrkA and p75 neurotrophin receptors regulate sympathetic neuron growth and target innervation.

In this report, we provide evidence that NGF and BDNF have functionally antagonistic actions on sympathetic neuron growth and target innervation, with NGF acting via TrkA to promote growth and BDNF via p75NTR to inhibit growth. Specifically, in cultured sympathetic neurons that themselves synthesize BDNF, exogenous BDNF inhibits and function-blocking BDNF antibodies enhance process outgrowth. Both exogenous and autocrine BDNF mediate this effect via p75NTR because (1) BDNF does not inhibit growth of neurons lacking p75NTR, (2) function-blocking p75NTR antibodies enhance NGF-mediated growth, and (3) p75NTR-/- sympathetic neurons grow more robustly in response to NGF than do their wild-type counterparts. To determine the physiological relevance of this functional antagonism, we examined the pineal gland, a well defined sympathetic target organ. BDNF is present in the pineal gland during target innervation, and incoming sympathetic axons are p75NTR positive. Moreover, the pineal glands of BDNF+/- and BDNF-/- mice are hyperinnervated with sympathetic fibers, and tyrosine hydroxylase (TH) levels are elevated. Increased tyrosine hydroxylase is also observed in the BDNF+/- carotid artery, another sympathetic neuron target. Thus, BDNF, made by sympathetic neurons and/or their target organs, acts via p75NTR to antagonize NGF-mediated growth and target innervation, suggesting that sympathetic target innervation is determined by the balance of positively and negatively acting neurotrophins present in developing and potentially mature targets.

Animals↗
PubMed1999

Early induction of Talpha1 alpha-tubulin transcription in neurons of the developing nervous system.

In this report, we have examined the relationship between the onset of neuronal gene transcription and neuronal development by characterizing expression of the early panneuronal Talpha1 alpha-tubulin promoter in developing neurons. In the peripheral nervous system, detectable expression of a beta-galactosidase transgene driven by the Talpha1 promoter (Talpha1:nlacZ) was coincident with neuronal birth dates, with the exception of sympathetic neuroblasts, which expressed the transgene prior to terminal mitosis. Similarly, in the central nervous system, the onset of beta-galactosidase expression was coincident with neuronal birth dates in most identifiable populations of central neurons. A small subpopulation of transgene-positive cells localized to ventricular zones, but the vast majority was observed in locations consistent with their identification as migrating and/or differentiating neurons. To determine more precisely the temporal relationship between transgene expression and terminal mitosis, we analyzed cultures of cortical progenitors that become postmitotic neurons in vitro. When initially plated, the vast majority of cells consisted of dividing, nestin-positive progenitors. Neurons differentiated from these progenitors as early as 1 day in vitro, as indicated by immunostaining for betaIII-tubulin, a neuron-specific tubulin isotype that is turned on shortly after terminal mitosis. Double-labeling studies showed that Talpha1:nlacZ expression was detectable in the same cells and at approximately the same time as was betaIII-tubulin, indicating that detectable transcription of the Talpha1 alpha-tubulin promoter commences at the time of terminal mitosis, at least in culture. This promoter, therefore, provides a valuable tool for genetic manipulation of early developing neurons in transgenic mice.

Animals↗
PubMed1999

Emergence of Schistosoma mansoni infection in upper Egypt: the Giza governorate.

We found an unexpectedly high prevalence of Schistosoma mansoni in a village in the Upper Egyptian governorate of Giza. Historically, S. mansoni is endemic in the northern Egyptian Nile Delta rather than in the southern Upper Egypt. This observation was made during an evaluation of a rural health care schistosomiasis surveillance program using a cross sectional survey for S. haematobium and S. mansoni in the village of El-Gezira El-Shakra El-Saf district in the Upper Egypt Giza Governorate. A 10% systematic random sample of households of the village was chosen. All persons in the selected houses were invited to submit urine and stool samples. All students from a primary school were also included in the study. Urine was screened by a polycarbonate filtration method and stool was examined using modified Kato-Katz technique. The prevalence of S. mansoni in the population sample and in the school children was 33.7% and 57.7%, respectively, whereas the prevalence of S. haematobium infection in the population sample and the school children was 7.4% and 10.6%, respectively. The prevalence of infection was highest in the younger age groups, and males were infected more than females. Review of Ministry of Health records showed that both species of vector snails, Bulinus truncatus and Biomphalaria alexandrina, were present from 1991 to 1995, and that B. alexandrina was more abundant than B. truncatus in the canals surrounding this village. The unexpected high prevalence of S. mansoni in this village indicates an urgent need to include training programs for S. mansoni surveillance in the primary health care facilities of Giza and to educate villagers to request examinations for S. mansoni as well as for S. haematobium infection.

Adolescent↗
PubMed1998

p53 is essential for developmental neuron death as regulated by the TrkA and p75 neurotrophin receptors.

Naturally occurring sympathetic neuron death is the result of two apoptotic signaling events: one normally suppressed by NGF/TrkA survival signals, and a second activated by the p75 neurotrophin receptor. Here we demonstrate that the p53 tumor suppressor protein, likely as induced by the MEKK-JNK pathway, is an essential component of both of these apoptotic signaling cascades. In cultured neonatal sympathetic neurons, p53 protein levels are elevated in response to both NGF withdrawal and p75NTR activation. NGF withdrawal also results in elevation of a known p53 target, the apoptotic protein Bax. Functional ablation of p53 using the adenovirus E1B55K protein inhibits neuronal apoptosis as induced by either NGF withdrawal or p75 activation. Direct stimulation of the MEKK-JNK pathway using activated MEKK1 has similar effects; p53 and Bax are increased and the subsequent neuronal apoptosis can be rescued by E1B55K. Expression of p53 in sympathetic neurons indicates that p53 functions downstream of JNK and upstream of Bax. Finally, when p53 levels are reduced or absent in p53+/- or p53-/- mice, naturally occurring sympathetic neuron death is inhibited. Thus, p53 is an essential common component of two receptor-mediated signal transduction cascades that converge on the MEKK-JNK pathway to regulate the developmental death of sympathetic neurons.

Animals↗
PubMed1998

Autocrine hepatocyte growth factor provides a local mechanism for promoting axonal growth.

In this report, we describe a novel local mechanism necessary for optimal axonal growth that involves hepatocyte growth factor (HGF). Sympathetic neurons of the superior cervical ganglion coexpress bioactive HGF and its receptor, the Met tyrosine kinase, both in vivo and in vitro. Exogenous HGF selectively promotes the growth but not survival of cultured sympathetic neurons; the magnitude of this growth effect is similar to that observed with exogenous NGF. Conversely, HGF antibodies that inhibit endogenous HGF decrease sympathetic neuron growth but have no effect on survival. This autocrine HGF is required locally by sympathetic axons for optimal growth, as demonstrated using compartmented cultures. Thus, autocrine HGF provides a local, intrinsic mechanism for promoting neuronal growth without affecting survival, a role that may be essential during developmental axogenesis or after neuronal injury.

Animals↗
PubMed1998

Functional evidence that BDNF is an anterograde neuronal trophic factor in the CNS.

In this report, we have tested the hypothesis that brain-derived neurotrophic factor (BDNF) is an anterograde neurotrophic factor in the CNS and have focused on central noradrenergic neurons that synthesize BDNF. Double-label immunocytochemistry for BDNF and dopamine-beta-hydroxylase (DBH), a marker for noradrenergic neurons, demonstrated that BDNF is partially localized to noradrenergic nerve fibers and terminals in the adult rat brain. To test the functional importance of this anterograde BDNF, we analyzed transgenic mice carrying a DBH-BDNF minigene. Increased synthesis of BDNF in noradrenergic neurons of DBH-BDNF mice caused elevated TrkB tyrosine kinase activation throughout postnatal life in the neocortex, a noradrenergic target region. This afferently regulated increase in TrkB receptor activity led to long-lasting alterations in cortical morphology. To determine whether noradrenergic neuron-expressed BDNF also anterogradely regulated neuronal survival, we examined a second noradrenergic target, neonatal facial motoneurons. One week after axotomy, 72% of facial motoneurons were lost in control animals, whereas only 30-35% were lost in DBH-BDNF transgenic mice. Altogether, these results indicate that BDNF is anterogradely transported to fibers and terminals of noradrenergic neurons, that anterogradely secreted BDNF causes activation of TrkB in target regions, and that this secretion has functional consequences for target neuron survival and differentiation. This presynaptic secretion of BDNF may provide a cellular mechanism for modulating neural circuitry, in either the developing or mature nervous system.

Age Factors↗
PubMed1998

A critical temporal requirement for the retinoblastoma protein family during neuronal determination.

In this report, we have examined the requirement for the retinoblastoma (Rb) gene family in neuronal determination with a focus on the developing neocortex. To determine whether pRb is required for neuronal determination in vivo, we crossed the Rb-/- mice with transgenic mice expressing beta-galactosidase from the early, panneuronal Talpha1 alpha-tubulin promoter (Talpha1:nlacZ). In E12.5 Rb-/- embryos, the Talpha1:nlacZ transgene was robustly expressed throughout the developing nervous system. However, by E14. 5, there were perturbations in Talpha1:nlacZ expression throughout the nervous system, including deficits in the forebrain and retina. To more precisely define the temporal requirement for pRb in neuronal determination, we functionally ablated the pRb family in wild-type cortical progenitor cells that undergo the transition to postmitotic neurons in vitro by expression of a mutant adenovirus E1A protein. These studies revealed that induction of Talpha1:nlacZ did not require proteins of the pRb family. However, in their absence, determined, Talpha1:nlacZ-positive cortical neurons underwent apoptosis, presumably as a consequence of "mixed signals" deriving from their inability to undergo terminal mitosis. In contrast, when the pRb family was ablated in postmitotic cortical neurons, there was no effect on neuronal survival, nor did it cause the postmitotic neurons to reenter the cell cycle. Together, these studies define a critical temporal window of requirement for the pRb family; these proteins are not required for induction of neuronal gene expression or for the maintenance of postmitotic neurons, but are essential for determined neurons to exit the cell cycle and survive.

Animals↗
PubMed1998

The p75 neurotrophin receptor mediates neuronal apoptosis and is essential for naturally occurring sympathetic neuron death.

To determine whether the p75 neurotrophin receptor (p75NTR) plays a role in naturally occurring neuronal death, we examined neonatal sympathetic neurons that express both the TrkA tyrosine kinase receptor and p75NTR. When sympathetic neuron survival is maintained with low quantities of NGF or KCl, the neurotrophin brain-derived neurotrophic factor (BDNF), which does not activate Trk receptors on sympathetic neurons, causes neuronal apoptosis and increased phosphorylation of c-jun. Function-blocking antibody studies indicate that this apoptosis is due to BDNF-mediated activation of p75NTR. To determine the physiological relevance of these culture findings, we examined sympathetic neurons in BDNF-/- and p75NTR-/- mice. In BDNF-/- mice, sympathetic neuron number is increased relative to BDNF+/+ littermates, and in p75NTR-/- mice, the normal period of sympathetic neuron death does not occur, with neuronal attrition occurring later in life. This deficit in apoptosis is intrinsic to sympathetic neurons, since cultured p75NTR-/- neurons die more slowly than do their wild-type counterparts. Together, these data indicate that p75NTR can signal to mediate apoptosis, and that this mechanism is essential for naturally occurring sympathetic neuron death.

Animals↗
PubMed1998

A mass chemotherapy trial of praziquantel on Schistosoma haematobium endemicity in Upper Egypt.

We evaluated the impact of mass chemotherapy using praziquantel (40 mg/kg of body weight) on Schistosoma haematobium endemicity in a typical village in Giza, Upper Egypt. The entire village population of 988, determined by census, was included in the study. At baseline all villagers were screened for S. haematobium and offered treatment with praziquantel regardless of their infection status. Infection was determined by nucleopore filtration and egg counts were reported per 10 ml of urine. Information on compliance to treatment or refusal to participate was also obtained at baseline. One month after treatment, all those who were found infected at baseline were re-examined for infection and re-treated if found positive. Baseline was conducted in late fall just prior to the low winter transmission season. Approximately one year after baseline, the entire village was requested to participate in an annual follow-up urine examination and treatment. The results showed that the baseline prevalence was reduced by 83.6% from 23.1% to 3.8%. Geometric mean egg counts decreased four-fold from 12.4 to 3.1. There was a 60-fold reduction in the estimated population egg count from 10,006 to 167. There were only eight cases of reinfection and five incident cases. The typical age specific prevalence curve was flattened, showing that those in the childhood age groups benefited the greatest reductions in endemicity. Treatment noncompliance was 30%, for which half had justifiable reasons. The others simply refused treatment many of whom were nevertheless examined for infection. Mass chemotherapy was found to be a feasible and an effective method to reduce measures of S. haematobium endemicity in Egypt.

Adolescent↗
PubMed1997

Transgenic mice expressing the intracellular domain of the p75 neurotrophin receptor undergo neuronal apoptosis.

We have asked whether p75(NTR) may play a role in neuronal apoptosis by producing transgenic mice that express the p75(NTR) intracellular domain within peripheral and central neurons. These animals showed profound reductions in numbers of sympathetic and peripheral sensory neurons as well as cell loss in the neocortex, where there is normally little or no p75(NTR) expression. Developmental loss of facial motor neurons was not observed, but induced expression of the p75(NTR) intracellular domain within adult animals led to increased motor neuron death after axotomy. Biochemical analyses suggest that these effects were not attributable to a p75(NTR)-dependent reduction in trk activation but instead indicate that the p75(NTR) intracellular domain may act as a constitutive activator of signaling cascades that regulate apoptosis in both peripheral and central neurons.

Animals↗
PubMed1997

Transcriptional repression of the growth-associated T alpha1 alpha-tubulin gene by target contact.

In this report, we address the molecular mechanisms that regulate axonal growth by focusing on the gene for one of the major axonal cytoskeletal proteins, T alpha1 alpha-tubulin. During the developmental growth of sympathetic neurons, transcription of a beta-galactosidase transgene driven by the T alpha1 promoter (T alpha1:nlacZ) was high until the time of target innervation and neuronal maturation, when it decreased significantly. In mature animals, T alpha1:nlacZ transcription remained relatively low until target contact was experimentally disrupted; when facial motoneurons were axotomized, T alpha1:nlacZ transgene expression increased, was maximal for 1-7 days, and, if neurons regenerated and reinnervated their target musculature, returned to control levels by 49 days. In contrast, if regeneration and reestablishment of target contact were inhibited, transgene expression remained elevated. To determine whether this increased transcription was due to the loss of target contact or to axonal loss, we transected sympathetic neurons that project to the eye either close to or far from their cell bodies. In both cases, when target contact was severed, T alpha1:nlacZ transcription increased. These experiments indicate that transcription of the T alpha1 alpha-tubulin promoter is repressed by target contact in both developing and mature neurons. We suggest that this repression is due to a target-derived "stop-growth" factor that retrogradely signals to regulate transcription of this and other genes that are required for axonal growth.

Animals↗
PubMed1997

Detection of brain-derived neurotrophic factor in a vesicular fraction of brain synaptosomes.

The mRNA encoding brain-derived neurotrophic factor (BDNF) is widely distributed in central nervous system neurons, including in hippocampus and cortex. However, little is known about the physiology of BDNF protein within neurons, including how it is processed or packaged and the mechanisms that control its release. In this study, we have used antibodies to monitor the subcellular distribution of BDNF in cortical extracts from adult rats treated with kainic acid. BDNF immunoreactivity is elevated in rat cortex 12 h after kainic acid treatment. The protein is enriched in a vesicular fraction isolated from lysed synaptosomes, its distribution being similar to that of synaptotagmin, which is associated with synaptic vesicles and large dense core vesicles at nerve terminals. The vesicular pool of BDNF is digested by proteinase K only in the presence of Triton X-100 suggesting localization of BDNF in membrane fractions. Immunocytochemistry detects diffuse and punctate BDNF staining within cell bodies and processes of cortical neurons from kainic acid-treated rats, as well as in mossy fiber terminals of rat hippocampus. Taken together, these data show that BDNF can accumulate axonally within a vesicular compartment of brain neurons. Results support the idea that endogenous BDNF may be transported anterogradely and released by regulated secretory mechanisms.

Animals↗
PubMed1997

Spatial regulation of neuronal gene expression in response to nerve growth factor.

To examine the cellular mechanisms whereby distally derived growth factors regulate nuclear responses in neurons, we have utilized compartmented cultures of sympathetic neurons to examine the regulation of two nerve growth factor (NGF)-inducible genes, tyrosine hydroxylase (TH) and p75 neurotrophin receptor (p75NTR). These studies demonstrate that NGF can signal retrogradely to mediate the induction of TH and p75NTR mRNAs. However, quantitative differences occurred as a function of the spatial localization of NGF exposure; application of NGF to cell bodies and proximal axons elicited peak levels of neuronal gene expression that were two- to threefold higher than when NGF was applied to distal axons alone. Furthermore, neurons responding maximally to NGF on distal axons were still able to respond to NGF administered to cell bodies and proximal axons. Biochemical analysis indicated that this difference in responsiveness was not due to differences in the number of TrkA/NGF receptors in the two compartments. Thus, although NGF signals retrogradely to mediate nuclear responses, the magnitude of these responses differs as a function of the spatial location of the activated NGF receptor:ligand complex. Moreover, these data suggest that neurons may be able to respond to a second cellular source of neurotrophins, even when target-derived neurotrophins are not limiting.

Animals↗
PubMed1997

NGF and neurotrophin-3 both activate TrkA on sympathetic neurons but differentially regulate survival and neuritogenesis.

In this report we examine the biological and molecular basis of the control of sympathetic neuron differentiation and survival by NGF and neurotrophin-3 (NT-3). NT-3 is as efficient as NGF in mediating neuritogenesis and expression of growth-associated genes in NGF-dependent sympathetic neurons, but it is 20-40-fold less efficient in supporting their survival. Both NT-3 and NGF induce similar sustained, long-term activation of TrkA, while NGF is 10-fold more efficient than NT-3 in mediating acute, short-term TrkA activity. At similar acute levels of TrkA activation, NT-3 still mediates neuronal survival two- to threefold less well than NGF. However, a mutant NT-3 that activates TrkC, but not TrkA, is unable to support sympathetic neuron survival or neuritogenesis, indicating that NT-3-mediated TrkA activation is necessary for both of these responses. On the basis of these data, we suggest that NGF and NT-3 differentially regulate the TrkA receptor both with regard to activation time course and downstream targets, leading to selective regulation of neuritogenesis and survival. Such differential responsiveness to two ligands acting through the same Trk receptor has important implications for neurotrophin function throughout the nervous system.

Animals↗
PubMed1997

Synaptic innervation density is regulated by neuron-derived BDNF.

In this report, we have examined the role of neuron-derived BDNF at an accessible synapse, that of preganglionic neurons onto their sympathetic neuron targets. Developing and mature sympathetic neurons synthesize BDNF, and preganglionic neurons express the full-length BDNF/TrkB receptor. When sympathetic neuron-derived BDNF is increased 2- to 4-fold in transgenic mice, preganglionic cell bodies and axons hypertrophy, and the synaptic innervation to sympathetic neurons is increased. Conversely, when BDNF synthesis is eliminated in BDNF -/- mice, preganglionic synaptic innervation to sympathetic neurons is decreased. Together these results indicate that variations in neuronal neurotrophin synthesis directly regulate neuronal circuitry by selectively modulating synaptic innervation density.

Animals↗