Measurement of axial rigidity and postural instability using wearable sensors

Axial Bradykinesia is an important feature of advanced Parkinson\u27s disease (PD). The purpose of this study is to quantify axial bradykinesia using wearable sensors with the long-term aim of quantifying these movements, while the subject performs routine domestic activities. We measured back movements during common daily activities such as pouring, pointing, walking straight and walking around a chair with a test system engaging a minimal number of Inertial Measurement (IM) based wearable sensors. Participants included controls and PD patients whose rotation and flexion of the back was captured by the time delay between motion signals from sensors attached to the upper and lower back. PD subjects could be distinguished from controls using only two sensors. These findings suggest that a small number of sensors and similar analyses could distinguish between variations in bradykinesia in subjects with measurements performed outside of the laboratory. The subjects could engage in routine activities leading to progressive assessments of therapeutic outcomes

An anatomical and single-cell gene expression characterisation of putative neurogenesis from nestin-expressing cells in the adult mouse midbrain

© 2016 Dr. Parisa FarzanehfarDue to significant midbrain dopamine (DA) cell loss in Parkinson’s disease (PD), it is generally believed the most effective and long-term treatment for PD motor symptoms will be midbrain DA cell replacement therapy, either by endogenous repair or cell transplantation. However, cell transplantation is hindered by failure of acquisition and maintenance of the DA phenotype by cells transplanted into the adult midbrain, and endogenous repair has not advanced very far because there appears to be very little or no neurogenesis and DA neurogenesis here. Evidence suggests Nestin-expressing neural precursor cells (NPCs) may give rise to neurones, including DA neurones in the adult midbrain, however this needs to be confirmed and underlying mechanisms established. The aims of my study were to: (1) Determine whether there is baseline neurogenesis and DA neurogenesis from Nestin+ cells in the adult mouse midbrain. (2) Investigate the gene expression profile of Nestin+ cells during different stages of putative ontogenesis in the adult mouse midbrain. (3) Test the possibility that putative neurogenesis and DA neurogenesis from Nestin+ cells in the adult mouse midbrain can be regulated by exogenous factors. To achieve these aims Nestin-expressing cells in the adult mouse midbrain were permanently labelled with enhanced yellow fluorescent protein (eYFP) by administering Tamoxifen to adult transgenic Nestin-CreERT2 x R26eYFP mice. Four-days to 8-months later eYFP+ cells were studied using a combination of whole-cell electrophysiology, single-cell qPCR and immunohistochemistry. eYFP+ cells expressed a range of genes consistent with birth, migration, neuronal and DA neuronal differentiation. Parsing the gene expression profiles of eYFP+ cells by their location in the midbrain indicated they arise anywhere throughout the midbrain and differentiate and integrate locally, rather than migrate long distances to populate midbrain nuclei. Most eYFP+ cells expressed mature neuronal genes, which was consistent also with their neuronal electrophysiology comprising action potentials and spontaneous post-synaptic currents. In comparison to neighbouring control (eYFP-) cells, eYFP+ cells expressed more immature neuronal genes (Pax6, Ngn2 & Msx1), which was also consistent with their more immature electrophysiology comprising hyper-excitability, hyperpolarized resting membrane potential, shorter duration sPSCs, and smaller membrane capacitance. Many (53%) eYFP+ cells expressed a combination of mature and immature neuronal genes at all time-points following Tamoxifen (an approximation of their age), including very ‘young’ cells (<2 months). Many (42%) of these young eYFP+ cells also had mature neuronal morphology (large cells with prominent processes) and electrophysiology. Nestin and Ki67 (a marker of cell division) were only expressed by large cells, and early on after Tamoxifen. Additionally, eYFP+ cells increased in number and size with time following Tamoxifen, and they decreased in number following administration of the anti-mitotic drug, Cytarabine (Ara-C). These data indicate that eYFP+ cells are capable of cell division. The level of expression of mature neuronal genes in eYFP+ cells also increased over a time-course of 7 months. In these respects, eYFP+ cells appear to proliferate, grow and differentiate into neurones, albeit slowly. To achieve the third aim of the study, bioinformatics analyses of the genes I identified as uniquely characterising eYFP+ cells (i.e. the proneuronal genes Pax6, Msx1 and Ngn2) highlighted valproic acid (VPA) as a drug that might regulate putative neurogenesis and DA neurogenesis from Nestin+ cells in the adult mouse midbrain. Infusion of VPA directly into the midbrain of adult mice for 2 weeks had no effect on the number of eYFP+ cells but significantly reduced the number of Pax6+, Pax6+/NeuN+ and eYFP+/NeuN+ cells. However, it also significantly reduced the number of NeuN+ cells generally (eYFP+ and eYFP-), pointing to a general effect of VPA on NeuN expression rather than neurogenesis from Nestin+ cells. Thus neither VPA nor targeting Pax6 appears to regulate of the number of eYFP+ cells or their differentiation into neurones in the adult mouse midbrain. In summary, my findings add to evidence of a small but significant population of Nestin-expressing cells in the adult mouse midbrain. While the origin of these cells remains unclear, my data suggest that: (1) some eYFP+ cells in the adult midbrain do undergo neurogenesis, albeit slowly; (2) Nestin expression in the adult midbrain is not limited to NPCs and classical neurogenesis, but also occurs in mature neurons, perhaps in relation to dendritic remodelling, cell death, or some other form of neural plasticity. 3) Neither VPA nor targeting Pax6 appears to regulate of the number of eYFP+ cells or their differentiation into neurones in the adult mouse midbrain. Further study of these cells may provide crucial information to assist DA cell replacement therapy for PD

Measurement of Axial Rigidity and Postural Instability Using Wearable Sensors

Axial Bradykinesia is an important feature of advanced Parkinson’s disease (PD). The purpose of this study is to quantify axial bradykinesia using wearable sensors with the long-term aim of quantifying these movements, while the subject performs routine domestic activities. We measured back movements during common daily activities such as pouring, pointing, walking straight and walking around a chair with a test system engaging a minimal number of Inertial Measurement (IM) based wearable sensors. Participants included controls and PD patients whose rotation and flexion of the back was captured by the time delay between motion signals from sensors attached to the upper and lower back. PD subjects could be distinguished from controls using only two sensors. These findings suggest that a small number of sensors and similar analyses could distinguish between variations in bradykinesia in subjects with measurements performed outside of the laboratory. The subjects could engage in routine activities leading to progressive assessments of therapeutic outcomes

Electrophysiological and gene expression characterization of the ontogeny of nestin-expressing cells in the adult mouse midbrain

The birth of new neurons, or neurogenesis, in the adult midbrain is important for progressing dopamine cell-replacement therapies for Parkinson's disease. Most studies suggest newborn cells remain undifferentiated or differentiate into glia within the adult midbrain. However, some studies suggest nestin + neural precursor cells (NPCs) have a propensity to generate new neurons here. We sought to confirm this by administering tamoxifen to adult NesCreERT2/R26eYFP transgenic mice, which permanently labelled adult nestin-expressing cells and their progeny with enhanced yellow fluorescent protein (eYFP). eYFP+ midbrain cells were then characterized 1–32 weeks later in acutely prepared brain slices using whole-cell patch clamp electrophysiology combined with single-cell RT-qPCR. Most eYFP+ cells exhibited a mature neuronal phenotype with large amplitude fast action potentials (APs), spontaneous post-synaptic currents (sPSCs), and expression of ‘mature’ neuronal genes (NeuN, Gad1, Gad2 and/or VGLUT2). This was the case even at the earliest time-point following tamoxifen (i.e. 1 week). In comparison to neighboring eYFP− (control) cells, eYFP+ cells discharged more APs per unit current injection, and had faster AP time-to-peak, hyperpolarized resting membrane potential, smaller membrane capacitance and shorter duration sPSCs. eYFP+ cells were also differentiated from eYFP− cells by increased expression of ‘immature’ pro-neuronal genes (Pax6, Ngn2 and/or Msx1). However, further analyses failed to reveal evidence of a place of birth, neuronal differentiation, maturation and integration indicative of classical neurogenesis. Thus our findings do not support the notion that nestin + NPCs in the adult SNc and midbrain generate new neurons via classical neurogenesis. Rather, they raise the possibility that mature neurons express nestin under unknown circumstances, and that this is associated with altered physiology and gene expression

Evidence of functional duplicity of Nestin expression in the adult mouse midbrain

Whether or not neurogenesis occurs in the adult substantia nigra pars compacta (SNc) is an important question relevant for developing better treatments for the motor symptoms of Parkinson's disease (PD). Although controversial, it is generally believed that dividing cells here remain undifferentiated or differentiate into glia, not neurons. However, there is a suggestion that Nestin-expressing neural precursor cells (NPCs) in the adult SNc have a propensity to differentiate into neurons, which we sought to confirm in the present study. Adult (>8-weeks old) transgenic NesCreERT2/GtROSA or NesCreERT2/R26eYFP mice were used to permanently label Nestin-expressing cells and their progeny with β-galactosidase (β-gal) or enhanced yellow fluorescent protein (eYFP), respectively. Most β-gal+ or eYFP+ cells were found in the ependymal lining of the midbrain aqueduct (Aq) and in the midline ventral to Aq. Smaller but significant numbers were in the periaqueductal gray (PAG), the ventral tegmental area (VTA), and in SNc. Low-level basal proliferation was evidenced by a modest increase in number of β-gal+ or eYFP+ cells over time, fewer β-gal+ or eYFP+ cells when mice were administered the anti-mitotic agent Cytarabine, and incorporation of the proliferation marker bromodeoxyuridine (BrdU) in a very small number of β-gal+ cells. No evidence of migration was found, including no immunoreactivity against the migration markers doublecortin (DCX) or polysialic acid neural cell adhesion molecule (PSA-NCAM), and no dispersal of β-gal+ or eYFP+ cells through the midbrain parenchyma over time. However, β-gal+ or eYFP+ cells did increase in size and express higher levels of mature neuronal genes over time, indicating growth and neuronal differentiation. In mice whose SNc dopamine neurons had been depleted with 6-hydroxy-dopamine, a model of PD, there were ~2-fold more β-gal+ cells in SNc specifically, although the proportion that were also NeuN+ was not affected. Remarkably, as early as 4 days following putative Nestin-expression, many β-gal+ or eYFP+ cells had mature neuronal morphology and were NeuN+. Furthermore, mature neuronal β-gal+ cells were immunoreactive against the self-renewal or pluripotency marker sex determining region Y-box 2 (Sox2). Overall, our data support the notion that some Nestin-expressing, presumably NPCs, have a limited capacity for proliferation, no capacity for migration, and a propensity to generate new neurons within the microenvironment of the adult midbrain. However, our data also suggest that significant numbers of extant midbrain neurons express Nestin and other classical neurogenesis markers in contexts that are presumably not neurogenic. These findings foreshadow duplicitous roles for Nestin and other molecules that are traditionally associated with neurogenesis in the adult midbrain, which should be considered in future PD research