BackgroundParaspeckles are subnuclear bodies assembled on a long non-coding RNA (lncRNA) NEAT1. Their enhanced formation in spinal neurons of sporadic amyotrophic lateral sclerosis (ALS) patients has been reported but underlying mechanisms are unknown. The majority of ALS cases are characterized by TDP-43 proteinopathy. In current study we aimed to establish whether and how TDP-43 pathology may augment paraspeckle assembly.MethodsParaspeckle formation in human samples was analysed by RNA-FISH and laser capture microdissection followed by qRT-PCR. Mechanistic studies were performed in stable cell lines, mouse primary neurons and human embryonic stem cell-derived neurons. Loss and gain of function for TDP-43 and other microRNA pathway factors were modelled by siRNA-mediated knockdown and protein overexpression.ResultsWe show that de novo paraspeckle assembly in spinal neurons and glial cells is a hallmark of both sporadic and familial ALS with TDP-43 pathology. Mechanistically, loss of TDP-43 but not its cytoplasmic accumulation or aggregation augments paraspeckle assembly in cultured cells. TDP-43 is a component of the microRNA machinery, and recently, paraspeckles have been shown to regulate pri-miRNA processing. Consistently, downregulation of core protein components of the miRNA pathway also promotes paraspeckle assembly. In addition, depletion of these proteins or TDP-43 results in accumulation of endogenous dsRNA and activation of type I interferon response which also stimulates paraspeckle formation. We demonstrate that human or mouse neurons in vitro lack paraspeckles, but a synthetic dsRNA is able to trigger their de novo formation. Finally, paraspeckles are protective in cells with compromised microRNA/dsRNA metabolism, and their assembly can be promoted by a small-molecule microRNA enhancer.ConclusionsOur study establishes possible mechanisms behind paraspeckle hyper-assembly in ALS and suggests their utility as therapeutic targets in ALS and other diseases with abnormal metabolism of microRNA and dsRNA.Electronic supplementary materialThe online version of this article (10.1186/s13024-018-0263-7) contains supplementary material, which is available to authorized users.

Efficient blood supply to the brain is of paramount importance to its normal functioning and improper blood flow can result in potentially devastating neurological consequences. Cerebral blood flow in response to neural activity is intrinsically regulated by a complex interplay between various cell types within the brain in a relationship termed neurovascular coupling. The breakdown of neurovascular coupling is evident across a wide variety of both neurological and psychiatric disorders including Alzheimer’s disease. Atherosclerosis is a chronic syndrome affecting the integrity and function of major blood vessels including those that supply the brain, and it is therefore hypothesised that atherosclerosis impairs cerebral blood flow and neurovascular coupling leading to cerebrovascular dysfunction. This review will discuss the mechanisms of neurovascular coupling in health and disease and how atherosclerosis can potentially cause cerebrovascular dysfunction that may lead to cognitive decline as well as stroke. Understanding the mechanisms of neurovascular coupling in health and disease may enable us to develop potential therapies to prevent the breakdown of neurovascular coupling in the treatment of vascular brain diseases including vascular dementia, Alzheimer’s disease and stroke.

Inhibitory interneurons can evoke vasodilation and vasoconstriction, making them potential cellular drivers of neurovascular coupling. However, the specific regulatory roles played by particular interneuron subpopulations remain unclear. Our purpose was therefore to adopt a cell-specific optogenetic approach to investigate how somatostatin (SST) and neuronal nitric oxide synthase (nNOS)-expressing interneurons might influence the neurovascular relationship. In mice, specific activation of SST- or nNOS-interneurons was sufficient to evoke hemodynamic changes. In the case of nNOS-interneurons, robust hemodynamic changes occurred with minimal changes in neural activity, suggesting that the ability of blood oxygen level dependent functional magnetic resonance imaging (BOLD fMRI) to reliably reflect changes in neuronal activity may be dependent on type of neuron recruited. Conversely, activation of SST-interneurons produced robust changes in evoked neural activity with shallow cortical excitation and pronounced deep layer cortical inhibition. Prolonged activation of SST-interneurons often resulted in an increase in blood volume in the centrally activated area with an accompanying decrease in blood volume in the surrounding brain regions, analogous to the negative BOLD signal. These results demonstrate the role of specific populations of cortical interneurons in the active control of neurovascular function.

20Inhibitory interneurons can evoke vasodilation and vasoconstriction, making them 21 potential cellular drivers of neurovascular coupling. However, the specific regulatory 22 roles played by particular interneuron subpopulations remain unclear. Our purpose 23 was therefore to adopt a cell-specific optogenetic approach to investigate how 24 somatostatin (SST) and neuronal nitric oxide synthase (NOS1)-expressing 25 interneurons might influence neurovascular relationships. In mice, specific activation 26 of SST-or NOS1-interneurons was sufficient to evoke haemodynamic changes similar 27 to those evoked by physiological whisker stimulation. In the case of NOS1-28 interneurons, robust haemodynamic changes occurred with minimal changes in neural 29 activity. Conversely, activation of SST-interneurons produced robust changes in 30 evoked neural activity with shallow cortical excitation and pronounced deep layer 31 cortical inhibition. This often resulted in a central increase in blood volume with 32 corresponding surround decrease, analogous to the negative BOLD signal. These 33 results demonstrate the role of specific populations of cortical interneurons in the 34 active control of neurovascular function.35 36 80 of cortical GABAergic interneurons have specific roles in NVC. Also, that the ability of 81 BOLD signals to act as a surrogate measure of local neural activation may in part be 82 dependent upon which subpopulation of neurons are being activated.83 84 4 Results 85 Short duration optogenetic stimulation of specific interneurons evokes a 86 localised haemodynamic response 87 Genetically modified mice expressing channelrhodopsin-2 (ChR2) in either SST-or 88 NOS1-expressing interneurons (referred to as SST-ChR2 or NOS1-ChR2 mice, 89 respectively) were used to investigate how light induced activity of these inhibitory 90 interneurons may alter cortical haemodynamics. Using an anaesthetised mouse 91 (Figure 1), we assessed whether short duration optogenetic stimulation of specific 92 subtypes of interneuron evoked a localised haemodynamic response, comparable to 93 that evoked by a mild physiological stimulus (mechanical whisker stimulation). 2-94 dimensional optical imaging spectroscopy (2D-OIS) was used to record high-95 resolution 2D maps of the changes in blood volume (Hbt), oxygenated haemoglobin 96 (HbO2) and reduced haemoglobin (Hbr) evoked by stimulation. Each animal initially 97 received a mechanical whisker stimulation (2s, 5Hz), evoking changes in Hbt, HbO2 98 and Hbr which were localised to the whisker barrel cortex (Figure 2A). These 99 haemodynamic changes allowed us to map the whisker barrel cortex and, in turn, 100 guide the placement of the optical fibre used for photostimulation (Figure 1). The time 101 series of the haemodynamic response to whisker stimulation shows an increase in Hbt 102and HbO2 during the stimulation with a corresponding washout of Hbr (Figure 2A). 103 5 104 A fibre-coupled blue (470nm) LED, placed directly above the whisker barrel cortex, 105 was used to apply photostimulat...

Impaired neurovascular coupling has been suggested as an early pathogenic factor in Alzheimer’s disease (AD), which could serve as an early biomarker of cerebral pathology. We have established an anaesthetic regime to allow repeated measurements of neurovascular function over three months in the J20 mouse model of AD (J20-AD) and wild-type (WT) controls. Animals were 9–12 months old at the start of the experiment. Mice were chronically prepared with a cranial window through which 2-Dimensional optical imaging spectroscopy (2D-OIS) was used to generate functional maps of the cerebral blood volume and saturation changes evoked by whisker stimulation and vascular reactivity challenges. Unexpectedly, the hemodynamic responses were largely preserved in the J20-AD group. This result failed to confirm previous investigations using the J20-AD model. However, a final acute electrophysiology and 2D-OIS experiment was performed to measure both neural and hemodynamic responses concurrently. In this experiment, previously reported deficits in neurovascular coupling in the J20-AD model were observed. This suggests that J20-AD mice may be more susceptible to the physiologically stressing conditions of an acute experimental procedure compared to WT animals. These results therefore highlight the importance of experimental procedure when determining the characteristics of animal models of human disease.