The Input-Output Equivalent Sources Method for Fast Simulations of Distributed Nonlinearities in Bulk Acoustic Wave Resonators and Filters.

This work presents a new method to analyze weak distributed nonlinear (NL) effects, with a focus on the generation of harmonics (H) and intermodulation products (IMD) in bulk acoustic wave (BAW) resonators and filters composed of them. The method consists of finding equivalent current sources [input-output equivalent sources (IOES)] at the H or IMD frequencies of interest that are applied to the boundary nodes of any layer that can contribute to the nonlinearities according to its local NL constitutive equations. The new methodology is compared with the harmonic balance (HB) analysis, by means of a commercial tool, of a discretized NL Mason model, which is the most used model for NL BAW resonators. While the computation time is drastically reduced, the results are fully identical. For the simulation of a seventh-order filter, the IOES method is around 700 times faster than the HB simulations.

Electro-thermo-mechanical model for bulk acoustic wave resonators.

We present the electro-thermo-mechanical constitutive relations, expanded up to the third order, for a BAW resonator. The relations obtained are implemented into a circuit model, which is validated with extensive linear and nonlinear measurements. The mathematical analysis, along with the modeling, allows us to identify the dominant terms, which are the material temperature derivatives and two intrinsic nonlinear terms, and explain, for the first time, all observable effects in a BAW resonator by use of a unified physical description. Moreover, the terms that are responsible for the second-harmonic generation and the frequency shift with dc voltage are shown to be the same.

Porous silicon bulk acoustic wave resonator with integrated transducer.

: We report that porous silicon acoustic Bragg reflectors and AlN-based transducers can be successfully combined and processed in a commercial solidly mounted resonator production line. The resulting device takes advantage of the unique acoustic properties of porous silicon in order to form a monolithically integrated bulk acoustic wave resonator.

Impaired adult olfactory bulb neurogenesis in the R6/2 mouse model of Huntington's disease.

BACKGROUND: Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder linked to expanded CAG-triplet nucleotide repeats within the huntingtin gene. Intracellular huntingtin aggregates are present in neurons of distinct brain areas, among them regions of adult neurogenesis including the hippocampus and the subventricular zone/olfactory bulb system. Previously, reduced hippocampal neurogenesis has been detected in transgenic rodent models of HD. Therefore, we hypothesized that mutant huntingtin also affects newly generated neurons derived from the subventricular zone of adult R6/2 HD mice. RESULTS: We observed a redirection of immature neuroblasts towards the striatum, however failed to detect new mature neurons. We further analyzed adult neurogenesis in the granular cell layer and the glomerular layer of the olfactory bulb, the physiological target region of subventricular zone-derived neuroblasts. Using bromodeoxyuridine to label proliferating cells, we observed in both neurogenic regions of the olfactory bulb a reduction in newly generated neurons. CONCLUSION: These findings suggest that the striatal environment, severely affected in R6/2 mice, is capable of attracting neuroblasts, however this region fails to provide sufficient signals for neuronal maturation. Moreover, in transgenic R6/2 animals, the hostile huntingtin-associated microenvironment in the olfactory bulb interferes with the survival and integration of new mature neurons. Taken together, endogenous cell repair strategies in HD may require additional factors for the differentiation and survival of newly generated neurons both in neurogenic and non-neurogenic regions.

Smad7 regulates the adult neural stem/progenitor cell pool in a transforming growth factor beta- and bone morphogenetic protein-independent manner.

Members of the transforming growth factor beta (TGF-beta) family of proteins modulate the proliferation, differentiation, and survival of many different cell types. Neural stem and progenitor cells (NPCs) in the adult brain are inhibited in their proliferation by TGF-beta and by bone morphogenetic proteins (BMPs). Here, we investigated neurogenesis in a hypomorphic mouse model for the TGF-beta and BMP inhibitor Smad7, with the hypothesis that NPC proliferation might be reduced due to increased TGF-beta and BMP signaling. Unexpectedly, we found enhanced NPC proliferation as well as an increased number of label-retaining cells in vivo. The enhanced proliferation potential of mutant cells was retained in vitro in neurosphere cultures. We observed a higher sphere-forming capacity as well as faster growth and cell cycle progression. Use of specific inhibitors revealed that these effects were independent of TGF-beta and BMP signaling. The enhanced proliferation might be at least partially mediated by elevated signaling via epidermal growth factor (EGF) receptor, as mutant cells showed higher expression and activation levels of the EGF receptor. Conversely, an EGF receptor inhibitor reduced the proliferation of these cells. Our data indicate that endogenous Smad7 regulates neural stem/progenitor cell proliferation in a TGF-beta- and BMP-independent manner.

Sputtered SiO2 as low acoustic impedance material for Bragg mirror fabrication in BAW resonators.

In this paper we describe the procedure to sputter low acoustic impedance SiO(2) films to be used as a low acoustic impedance layer in Bragg mirrors for BAW resonators. The composition and structure of the material are assessed through infrared absorption spectroscopy. The acoustic properties of the films (mass density and sound velocity) are assessed through X-ray reflectometry and picosecond acoustic spectroscopy. A second measurement of the sound velocity is achieved through the analysis of the longitudinal lambda/2 resonance that appears in these silicon oxide films when used as uppermost layer of an acoustic reflector placed under an AlN-based resonator.

Dopamine receptor activation promotes adult neurogenesis in an acute Parkinson model.

Cell proliferation of neural progenitors in the subventricular zone (SVZ) of Parkinson disease (PD) patients and animal models is decreased. It was previously demonstrated that the neurotransmitter dopamine modulates cell proliferation in the embryonic brain. The aim of the present study was to analyze whether oral treatment with the dopamine receptor agonist pramipexole (PPX) modulates adult neurogenesis in the SVZ/olfactory bulb system in a dopaminergic lesion model. 6-Hydroxydopamine (6-OHDA) lesioned adult rats received either PPX (1.0 mg/kg) or PBS orally twice daily and bromodeoxyuridine (BrdU, a cell proliferation marker) for 10 days and were perfused immediately after treatment or 4 weeks after PPX withdrawal. Stereological analysis revealed a significant augmentation in SVZ proliferation by PPX. Consecutively, enhanced neuronal differentiation and more new neurons were present in the olfactory bulb 4 weeks after PPX withdrawal. In addition, dopaminergic neurogenesis was increased in the olfactory bulb after PPX treatment. Motor activity as assessed by using an open field paradigm was permanently increased even after long term PPX withdrawal. In addition, we demonstrate that D2 and D3 receptors are present on adult rat SVZ-derived neural progenitors in vitro, and PPX specifically increased mRNA levels of epidermal growth factor receptor (EGF-R) and paired box gene 6 (Pax6). Oral PPX treatment selectively increases adult neurogenesis in the SVZ-olfactory bulb system by increasing proliferation and cell survival of newly generated neurons. Analyzing the neurogenic fate decisions mediated by D2/D3 signaling pathways may lead to new avenues to induce neural repair in the adult brain.

Changes in adult olfactory bulb neurogenesis in mice expressing the A30P mutant form of alpha-synuclein.

In familial and sporadic forms of Parkinson's disease (PD), alpha-synuclein pathology is present in the brain stem nuclei and olfactory bulb (OB) long before Lewy bodies are detected in the substantia nigra. The OB is an active region of adult neurogenesis, where newly generated neurons physiologically integrate. While accumulation of wild-type alpha-synuclein is one of the pathogenic hallmarks of non-genetic forms of PD, the A30P alpha-synuclein mutation results in an earlier disease onset and a severe clinical phenotype. Here, we study the regulation of adult neurogenesis in the subventricular zone (SVZ)/OB system in a tetracycline-suppressive (tet-off) transgenic model of synucleinopathies, expressing human mutant A30P alpha-synuclein under the control of the calcium/calmodulin-dependent protein kinase II alpha (CaMK) promoter. In A30P transgenic mice alpha-synuclein was abundant at the site of integration in the glomerular cell layer of the OB. Without changes in proliferation in the SVZ, significantly fewer newly generated neurons were observed in the OB granule cell and glomerular layers of A30P transgenic mice than in controls, most probably due to increased cell death. By tetracycline-dependent abrogation of A30P alpha-synuclein expression, OB neurogenesis and programmed cell death was restored to control levels. Our results indicate that, using A30P conditional (tet-off) mice, A30P alpha-synuclein has a negative impact on olfactory neurogenesis and suppression of A30P alpha-synuclein enhances survival of newly generated neurons. This finding suggests that interfering with alpha-synuclein pathology can rescue newly generated neurons, possibly leading to new targets for therapeutic interventions in synucleinopathies.

In vivo optical imaging of neurogenesis: watching new neurons in the intact brain.

Adult neurogenesis is a highly dynamic process modulated by several pathologic and environmental factors, as well as by various compounds. So far, available techniques to study neurogenesis are lengthy and personnel and cost intensive. We developed a new tool based on the doublecortin promoter driving the expression of the luciferase reporter gene (DCX-promo-luciferase) in transgenic mice to perform in vivo imaging of neurogenesis. Indeed, the DCX-promo-luciferase mice allowed optical in vivo imaging of the onset of and increase in neurogenesis in developing fetal brains, as well as imaging of neurogenesis in the intact adult mouse central nervous system. Moreover, the capacity to specifically detect a small number of migrating neuronal precursors in vivo after transplantation is for the first time feasible using this DCX-promo-luciferase transgenic tool. The present imaging approach offers several crucial advantages over methods currently available, such as bromodeoxyuridine incorporation or labeling using iron oxide nanoparticles. Hence, it allows longitudinal study of neurogenesis in intact animals without the requirement of cellular prelabeling. Moreover, it guarantees that detection is specific for neuronal precursors and restricted to viable cells. Hence, our DCX-promo-luciferase transgenic model constitutes an effective tool that answers the pressing need for rapid investigation of the impact on neurogenesis of a large number of candidate compounds waiting to be tested.

Dopaminergic lesion enhances growth factor-induced striatal neuroblast migration.

Adult neurogenesis persists in the subventricular zone and is decreased in Parkinson disease (PD). The therapeutic potential of neurogenesis in PD requires understanding of mechanisms of 1) neural stem cell generation; 2) their guidance to the lesion site; and 3) the environment that enables neuronal differentiation, survival, and functional integration. We examined the combined intraventricular infusion of epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF-2) in a 6-hydroxydopamine-induced rodent model of PD. Epidermal growth factor and FGF-2 induced a massive increase in cell proliferation and in numbers of doublecortin-expressing neuroblasts in the subventricular zone. These growth factors also increased dopaminergic neurogenesis in the olfactory bulb and promoted the migration of newly generated neuroblasts from the subventricular zone into the adjacent striatum. The effects of EGF and FGF-2 were present in unlesioned animals but were dramatically enhanced in 6-hydroxydopamine-lesioned animals. These findings suggest that newly generated neuroblasts may be redirected to the region of dopaminergic deficit, and that EGF and FGF-2 can enhance dopaminergic neurogenesis in the olfactory bulb but not in the striatum. Similar mechanisms may be involved in the increased numbers of dopaminergic neurons observed in the olfactory bulbs of PD patients and their functional olfactory deficits.

Mutant alpha-synuclein exacerbates age-related decrease of neurogenesis.

In Parkinson disease, wild-type alpha-synuclein accumulates during aging, whereas alpha-synuclein mutations lead to an early onset and accelerated course of the disease. The generation of new neurons is decreased in regions of neurogenesis in adult mice overexpressing wild-type human alpha-synuclein. We examined the subventricular zone/olfactory bulb neurogenesis in aged mice expressing either wild-type human or A53T mutant alpha-synuclein. Aging wild-type and mutant alpha-synuclein-expressing animals generated significantly fewer new neurons than their non-transgenic littermates. This decreased neurogenesis was caused by a reduction in cell proliferation within the subventricular zone of mutant alpha-synuclein mice. In contrast, no difference was detected in mice overexpressing the wild-type allele. Also, more TUNEL-positive profiles were detected in the subventricular zone, following mutant alpha-synuclein expression and in the olfactory bulb, following wild-type and mutant alpha-synuclein expression. The impaired neurogenesis in the olfactory bulb of different transgenic alpha-synuclein mice during aging highlights the need to further explore the interplay between olfactory dysfunction and neurogenesis in Parkinson disease.

Physical activity fails to rescue hippocampal neurogenesis deficits in the R6/2 mouse model of Huntington's disease.

Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder linked to a mutation in the huntingtin gene leading to protein aggregation in neurons. The generation of new neurons in neurogenic regions, such as the subventricular zone of the lateral ventricle and the dentate gyrus of the hippocampus, is affected by these aggregation processes. In particular, hippocampal neurogenesis is reduced in the R6/2 transgenic mouse model of HD. Since physical activity stimulates adult hippocampal neurogenesis, we examined whether running is capable to rescue the impaired hippocampal neurogenesis in R6/2 mice. Proliferation of hippocampal cells measured by proliferating cell nuclear antigen (PCNA) marker was reduced in R6/2 animals by 64% compared to wild type mice. Accordingly, newly generated neurons labeled with doublecortin (DCX) were diminished by 60% in the hippocampus of R6/2 mice. Furthermore, the number of newly generated mature neurons was decreased by 76%. Within the hippocampus of wild type animals, a four-week running period resulted in a doubling of PCNA-, DCX-, and bromo-deoxyuridine (BrdU)-labeled cells. However, physical exercise failed to stimulate proliferation and survival of newly generated neurons in R6/2 transgenic mouse model of HD. These findings suggest that mutant huntingtin alters the hippocampal microenvironment thus resulting in an impaired neurogenesis. Importantly, this adverse microenvironment impeded neurogenesis upregulation such as induced by physical exercise. Future studies need to decipher the molecular pathways involved in repressing the generation of new neurons after physical activity in huntingtin transgenic rodents.

Targeted transgene expression in neuronal precursors: watching young neurons in the old brain.

Progress in the field of neurogenesis is limited by the lack of animal models allowing direct detection and analysis of living cells participating in neurogenesis. We engineered a transgenic mouse model that expresses the fluorescent reporter proteins enhanced green fluorescent protein or Discoma sp. reef coral red fluorescent protein under the control of the doublecortin (DCX) promoter, a gene specifically and transiently active in neuronal precursors and young neurons. The expression of the reporter proteins correlated with expression of the endogenous DCX protein, and with developmental and adult neurogenesis. Neurogenesis was unaffected by the presence of the fluorescent proteins. The transgenic mice allowed direct identification of the very few newly generated neurons present in the aged brain. We performed electrophysiological analysis and established that newly generated hippocampal granule cells in aged and young mice shared identical physiological properties. Hence, although the rate of neurogenesis tapers with ageing, a population of highly excitable young neurons indistinguishable to those found in younger animals is continuously generated. Therefore, maintenance of the fundamental properties of neuronal precursors even at advanced age suggests that stimulation of neurogenesis may constitute a valid strategy to counteract age-related neuronal loss and cognitive declines.

Transforming growth factor-beta1 is a negative modulator of adult neurogenesis.

Transforming growth factor (TGF)-beta1 has multiple functions in the adult central nervous system (CNS). It modulates inflammatory responses in the CNS and controls proliferation of microglia and astrocytes. In the diseased brain, TGF-beta1 expression is upregulated and, depending on the cellular context, its activity can be beneficial or detrimental regarding regeneration. We focus on the role of TGF-beta1 in adult neural stem cell biology and neurogenesis. In adult neural stem and progenitor cell cultures and after intracerebroventricular infusion, TGF-beta1 induced a long-lasting inhibition of neural stem and progenitor cell proliferation and a reduction in neurogenesis. In vitro, although TGF-beta1 specifically arrested neural stem and progenitor cells in the G0/1 phase of the cell cycle, it did not affect the self-renewal capacity and the differentiation fate of these cells. Also, in vivo, TGF-beta1 did not influence the differentiation fate of newly generated cells as shown by bromo-deoxyuridine incorporation experiments. Based on these data, we suggest that TGF-beta1 is an important signaling molecule involved in the control of neural stem and progenitor cell proliferation in the CNS. This might have potential implications for neurogenesis in a variety of TGF-beta1-associated CNS diseases and pathologic conditions.

Striatal deafferentation increases dopaminergic neurogenesis in the adult olfactory bulb.

Dopaminergic loss is known to be one of the major hallmarks of Parkinson disease (PD). In addition to its function as a neurotransmitter, dopamine plays significant roles in developmental and adult neurogenesis. Both dopaminergic deafferentation and stimulation modulate proliferation in the subventricular zone (SVZ)/olfactory bulb system as well as in the hippocampus. Here, we study the impact of 6-hydroxydopamine (6-OHDA) lesions to the medial forebrain bundle on proliferation and neuronal differentiation of newly generated cells in the SVZ/olfactory bulb axis in adult rats. Proliferation in the SVZ decreased significantly after dopaminergic deafferentation. However, the number of neural progenitor cells expressing the proneuronal cell fate determinant Pax-6 increased in the SVZ. Survival and quantitative cell fate analysis of newly generated cells revealed that 6-OHDA lesions induced opposite effects in the two different regions of neurogenesis in the olfactory bulb: a transient decrease in the granule cell layer contrasts to a sustained increase of newly generated neurons in the glomerular layer. These data point towards a shift in the ratio of newly generated interneurons in the olfactory bulb layers. Dopaminergic neurogenesis in the glomerular layer tripled after lesioning and consistent with this finding, the total number of tyrosine hydroxylase (TH)-positive cells increased. Thus, loss of dopaminergic input to the SVZ led to a distinct cell fate decision towards stimulation of dopaminergic neurogenesis in the olfactory bulb glomerular layer. This study supports the accumulating evidence that neurotransmitters play a crucial role in determining survival and differentiation of newly generated neurons.

Doublecortin expression levels in adult brain reflect neurogenesis.

Progress in the field of neurogenesis is currently limited by the lack of tools enabling fast and quantitative analysis of neurogenesis in the adult brain. Doublecortin (DCX) has recently been used as a marker for neurogenesis. However, it was not clear whether DCX could be used to assess modulations occurring in the rate of neurogenesis in the adult mammalian central nervous system following lesioning or stimulatory factors. Using two paradigms increasing neurogenesis levels (physical activity and epileptic seizures), we demonstrate that quantification of DCX-expressing cells allows for an accurate measurement of modulations in the rate of adult neurogenesis. Importantly, we excluded induction of DCX expression during physiological or reactive gliogenesis and excluded also DCX re-expression during regenerative axonal growth. Our data validate DCX as a reliable and specific marker that reflects levels of adult neurogenesis and its modulation. We demonstrate that DCX is a valuable alternative to techniques currently used to measure the levels of neurogenesis. Importantly, in contrast to conventional techniques, analysis of neurogenesis through the detection of DCX does not require in vivo labelling of proliferating cells, thereby opening new avenues for the study of human neurogenesis under normal and pathological conditions.

Human wild-type alpha-synuclein impairs neurogenesis.

Neurodegenerative diseases classified as synucleinopathies are characterized by alpha-synuclein inclusions. In these disorders, alpha-synuclein accumulates within glial or neuronal cells in the brain including regions of adult neurogenesis. We hypothesized a pathophysiological role for alpha-synuclein in newly generated cells of the adult brain and in this study examined regions of neurogenesis in adult mice overexpressing human wild-type alpha-synuclein under the control of the platelet-derived growth factor promoter. The number of proliferating cells and the fate of newly generated cells were analyzed in the olfactory bulb system and in the hippocampal dentate gyrus. There were no effects on proliferation detectable; however, significantly less neurogenesis and fewer neurons were observed in the olfactory bulb as well as in the hippocampus of adult human alpha-synuclein mice compared to control littermates. This effect was almost exclusively due to diminished survival of neuronal precursors in the target regions of neurogenesis. Our data imply that the finely tuned equilibrium of neuronal cell birth and death in neurogenic regions may be altered in human alpha-synuclein-overexpressing mice. We hypothesize that reduced adult neurogenesis in the olfactory bulb may contribute to olfactory deficits in neurodegenerative disorders associated with alpha-synuclein inclusions.

Long-term survival and cell death of newly generated neurons in the adult rat olfactory bulb.

In the adult rat olfactory bulb, neurons are continually generated from progenitors that reside in the lateral ventricle wall. This study investigates long-term survival and cell death of newly generated cells within the adult olfactory bulb. After injecting rats at 2 months of age with 5-bromodeoxyuridine (BrdU), the newly generated cells were quantified over a period of 19 months. A peak of BrdU-positive cells was reached in the olfactory bulb 1 month after BrdU injection, when all new cells have finished migrating from the ventricle wall. Thereafter, a reduction of BrdU-positive cells to about 50% was observed and it was confirmed by dUTP-nick end-labelling (TUNEL) that progenitors and young neurons undergo programmed cell death. However, cells that survived the first 3 months after BrdU injection persisted for up to 19 months. The majority of the BrdU-positive cells that reach the olfactory bulb differentiate into granule cells, but a small fraction migrate further into the glomerular layer. These newborn cells differentiate more slowly into periglomerular interneurons, with a delay of more than 1 month when compared to the granule cells. The newly generated periglomerular neurons, among them a significant fraction of dopaminergic cells, showed a similar decline in number compared to the granule cell layer and long-term survival for the remaining new neurons of up to 19 months. Rather than replacing old neurons, this data suggests that adult olfactory bulb neurogenesis utilizes the overproduction and turnover of young neurons, which is reminiscent of the cellular dynamics observed during brain development.