The Contribution of Calcium-Activated Potassium Channel Dysfunction to Altered Purkinje Neuron Membrane Excitability in Spinocerebellar Ataxia

Spinocerebellar ataxias (SCA) are a family of dominantly-inherited neurodegenerative disorders which affect movement and coordination. Patients experience the shared features of cerebellar ataxia, characterized by uncoordinated limb movements, unsteady gait, and difficulties with balance and posture. Although the underlying genetic causes of SCA are diverse, these diseases are often associated with degeneration of neurons within the cerebellum. Purkinje neurons, which are the sole output of the cerebellar cortex, show enhanced vulnerability to dysfunction and degeneration in ataxia. Therefore, therapies which target aberrant Purkinje neuron function have great potential for the treatment of cerebellar ataxia. In order to determine whether targeting Purkinje neuron dysfunction is a reasonable strategy to improve motor impairment in SCA, I sought to identify molecular targets which contribute to altered Purkinje neuron spiking in SCA. I performed studies in mouse models of SCA1 and SCA7, two of the polyglutamine (polyQ) SCAs, diseases which are caused by an expanded CAG repeat sequence in their respective disease-causing genes. In the polyQ SCAs, expansion of this CAG repeat sequence beyond a pathogenic length results in symptoms of ataxia. Previously, our laboratory and others have shown that alterations in Purkinje neuron spiking are present at the onset of motor impairment in polyQ SCA, suggesting that Purkinje neuron dysfunction may directly contribute to motor impairment. However, the molecular basis for this dysfunction remains unclear. In the present studies, I illustrate that there is shared transcriptional disruption present across multiple models of polyQ SCA, including SCA1 and SCA7. The resulting downregulated mRNA transcripts are highly enriched for genes related to Purkinje neuron excitability. Four of these genes, Kcnma1 (the large-conductance calcium-activated potassium channel, BK), Cacna1g (T-type voltage-gated calcium channel), Itpr1 (inositol trisphosphate receptor), and Trpc3 (transient receptor potential cation channel, type C3) form an excitability module of calcium sources and an effector calcium-activated potassium (KCa) channel. Through a series of experiments, I illustrate that pharmacologic blockade of both calcium sources and KCa channels is necessary to induce irregular spiking in wild-type Purkinje neurons and, importantly, genetic replacement of the effector KCa channel, BK, is sufficient to restore regular spiking to SCA7 Purkinje neurons even in the presence of reduced calcium availability. These studies suggest that KCa dysfunction through a disrupted calcium homeostasis module may be a molecular target in ataxia. Next, I sought to determine whether KCa channel dysfunction can be targeted to improve motor impairment in a mouse model of SCA1. I performed a targeted screen of KCa channel activating-compounds and other potassium channel-activating compounds, and identified a combination of FDA-approved compounds which improves alterations in both SCA1 Purkinje neuron spiking and motor performance in SCA1 mice. In a small tolerability study, these same compounds were tolerated by human SCA patients and appear to improve symptoms. These results argue for a future clinical trial of potassium channel activators to treat SCA. Together, these studies address the hypothesis that calcium-activated potassium channel dysfunction is central to Purkinje neuron electrophysiologic dysfunction and motor impairment in spinocerebellar ataxia, and that potassium channel-activating compounds are outstanding candidates for the improvement of motor function in spinocerebellar ataxia. The overall impact of these studies is to establish a link between potassium channel dysfunction and motor impairment as a general mechanism of spinocerebellar ataxia and to demonstrate that potassium channel activation merits consideration for the treatment of human ataxia.PHDMolecular and Integrative PhysiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/145935/1/dbushart_1.pd

Padding Ain't Enough: Assessing the Privacy Guarantees of Encrypted DNS

DNS over TLS (DoT) and DNS over HTTPS (DoH) encrypt DNS to guard user privacy by hiding DNS resolutions from passive adversaries. Yet, past attacks have shown that encrypted DNS is still sensitive to traffic analysis. As a consequence, RFC 8467 proposes to pad messages prior to encryption, which heavily reduces the characteristics of encrypted traffic. In this paper, we show that padding alone is insufficient to counter DNS traffic analysis. We propose a novel traffic analysis method that combines size and timing information to infer the websites a user visits purely based on encrypted and padded DNS traces. To this end, we model DNS sequences that capture the complexity of websites that usually trigger dozens of DNS resolutions instead of just a single DNS transaction. A closed world evaluation based on the Alexa top-10k websites reveals that attackers can deanonymize at least half of the test traces in 80.2% of all websites, and even correctly label all traces for 32.0% of the websites. Our findings undermine the privacy goals of state-of-the-art message padding strategies in DoT/DoH. We conclude by showing that successful mitigations to such attacks have to remove the entropy of inter-arrival timings between query responses

A universal relationship between magnetization and local structure changes below the ferromagnetic transition in La_{1-x}Ca_xMnO_3; evidence for magnetic dimers

We present extensive X-ray Absorption Fine Structure (XAFS) measurements on La_{1-x}Ca_xMnO_3 as a function of B-field (to 11T) and Ca concentration, x (21-45%). These results reveal local structure changes (associated with polaron formation) that depend only on the magnetization for a given sample, irrespective of whether the magnetization is achieved through a decrease in temperature or an applied magnetic field. Furthermore, the relationship between local structure and magnetization depends on the hole doping. A model is proposed in which a filamentary magnetization initially develops via the aggregation of pairs of Mn atoms involving a hole and an electron site. These pairs have little distortion and it is likely that they pre-form at temperatures above T_c.Comment: 5 pages, 5 figures (1 with 2 parts) -- v2. new data added (updated figures); discussion expande