Long-Term Pioglitazone Treatment Has No Significant Impact on Microglial Activation and Tau Pathology in P301S Mice.

Neuroinflammation is one disease hallmark on the road to neurodegeneration in primary tauopathies. Thus, immunomodulation might be a suitable treatment strategy to delay or even prevent the occurrence of symptoms and thus relieve the burden for patients and caregivers. In recent years, the peroxisome proliferator-activated receptor γ (PPARγ) has received increasing attention as it is immediately involved in the regulation of the immune system and can be targeted by the anti-diabetic drug pioglitazone. Previous studies have shown significant immunomodulation in amyloid-ß (Aß) mouse models by pioglitazone. In this study, we performed long-term treatment over six months in P301S mice as a tauopathy model with either pioglitazone or placebo. We performed serial 18 kDa translocator protein positron-emission-tomography (TSPO-PET) imaging and terminal immunohistochemistry to assess microglial activation during treatment. Tau pathology was quantified via immunohistochemistry at the end of the study. Long-term pioglitazone treatment had no significant effect on TSPO-PET, immunohistochemistry read-outs of microglial activation, or tau pathology levels in P301S mice. Thus, we conclude that pioglitazone modifies the time course of Aß-dependent microglial activation, but does not significantly modulate microglial activation in response to tau pathology.

A Non-Canonical Cortico-Amygdala Inhibitory Loop.

Discriminating between auditory signals of different affective value is critical for the survival and success of social interaction of an individual. Anatomical, electrophysiological, imaging, and optogenetics approaches have established that the auditory cortex (AC) by providing auditory information to the lateral amygdala (LA) via long-range excitatory glutamatergic projections has an impact on sound-driven aversive/fear behavior. Here we test the hypothesis that the LA also receives GABAergic projections from the cortex. We addressed this fundamental question by taking advantage of optogenetics, anatomical, and electrophysiology approaches and directly examining the functional effects of cortical GABAergic inputs to LA neurons of the mouse (male/female) AC. We found that the cortex, via cortico-lateral-amygdala somatostatin neurons (CLA-SOM), has a direct inhibitory influence on the output of the LA principal neurons. Our results define a CLA long-range inhibitory circuit (CLA-SOM inhibitory projections → LA principal neurons) underlying the control of spike timing/generation in LA and LA-AC projecting neurons, and attributes a specific function to a genetically defined type of cortical long-range GABAergic neurons in CLA communication.SIGNIFICANCE STATEMENT It is very well established that cortical auditory inputs to the lateral amygdala are exclusively excitatory and that cortico-amygdala neuronal activity has been shown to be involved in sound-driven aversive/fear behavior. Here, for the first time, we show that the lateral amygdala receives long-range GABAergic projection from the auditory cortex and these form direct monosynaptic inhibitory connections onto lateral amygdala principal neurons. Our results define a cellular basis for direct inhibitory communication from auditory cortex to the lateral amygdala, suggesting that the timing and ratio of excitation and inhibition, two opposing forces in the mammalian cerebral cortex, can dynamically affect the output of the lateral amygdala, providing a general mechanism for fear/aversive behavior driven by auditory stimuli.

A fully organic retinal prosthesis restores vision in a rat model of degenerative blindness.

The degeneration of photoreceptors in the retina is one of the major causes of adult blindness in humans. Unfortunately, no effective clinical treatments exist for the majority of retinal degenerative disorders. Here we report on the fabrication and functional validation of a fully organic prosthesis for long-term in vivo subretinal implantation in the eye of Royal College of Surgeons rats, a widely recognized model of retinitis pigmentosa. Electrophysiological and behavioural analyses reveal a prosthesis-dependent recovery of light sensitivity and visual acuity that persists up to 6-10 months after surgery. The rescue of the visual function is accompanied by an increase in the basal metabolic activity of the primary visual cortex, as demonstrated by positron emission tomography imaging. Our results highlight the possibility of developing a new generation of fully organic, highly biocompatible and functionally autonomous photovoltaic prostheses for subretinal implants to treat degenerative blindness.

Filamin A promotes dynamin-dependent internalization of hyperpolarization-activated cyclic nucleotide-gated type 1 (HCN1) channels and restricts Ih in hippocampal neurons.

The actin-binding protein filamin A (FLNa) regulates neuronal migration during development, yet its roles in the mature brain remain largely obscure. Here, we probed the effects of FLNa on the regulation of ion channels that influence neuronal properties. We focused on the HCN1 channels that conduct Ih, a hyperpolarization-activated current crucial for shaping intrinsic neuronal properties. Whereas regulation of HCN1 channels by FLNa has been observed in melanoma cell lines, its physiological relevance to neuronal function and the underlying cellular pathways that govern this regulation remain unknown. Using a combination of mutational, pharmacological, and imaging approaches, we find here that FLNa facilitates a selective and reversible dynamin-dependent internalization of HCN1 channels in HEK293 cells. This internalization is accompanied by a redistribution of HCN1 channels on the cell surface, by accumulation of the channels in endosomal compartments, and by reduced Ih density. In hippocampal neurons, expression of a truncated dominant-negative FLNa enhances the expression of native HCN1. Furthermore, acute abrogation of HCN1-FLNa interaction in neurons, with the use of decoy peptides that mimic the FLNa-binding domain of HCN1, abolishes the punctate distribution of HCN1 channels in neuronal cell bodies, augments endogenous Ih, and enhances the rebound-response ("voltage-sag") of the neuronal membrane to transient hyperpolarizing events. Together, these results support a major function of FLNa in modulating ion channel abundance and membrane trafficking in neurons, thereby shaping their biophysical properties and function.

Striatal dopamine neurotransmission is altered in age- and region-specific manner in a Parkinson's disease transgenic mouse.

Dopamine (DA) plays a critical role in striatal motor control. The drop in DA level within the dorsal striatum is directly associated with the appearance of motor symptoms in Parkinson's disease (PD). The progression of the disease and inherent disruption of the DA neurotransmission has been closely related to accumulation of the synaptic protein α-synuclein. However, it is still unclear how α-synuclein affects dopaminergic terminals in different areas of dorsal striatum. Here we demonstrate that the overexpression of human α-synuclein (h-α-syn) interferes with the striatal DA neurotransmission in an age-dependent manner, preferentially in the dorsolateral striatum (DLS) of PDGF-h-α-syn mice. While 3-month-old mice showed an increase at the onset of h-α-syn accumulation in the DLS, 12-month-old mice revealed a decrease in electrically-evoked DA release. The enhanced DA release in 3-month-old mice coincided with better performance in a behavioural task. Notably, DA amplitude alterations were also accompanied by a delay in the DA clearance independently from the animal age. Structurally, dopamine transporter (DAT) was found to be redistributed in larger DAT-positive clumps only in the DLS of 3- and 12-month-old mice. Together, our data provide new insight into the vulnerability of DLS and suggest DAT-related dysfunctionalities from the very early stages of h-α-syn accumulation.

Photosensitized and Photothermal Stimulation of Cellular Membranes by Organic Thin Films and Nanoparticles.

Conjugated polymers are increasingly exploited for biomedical applications. In this work, we explored the optical characteristics of conjugated polymers of variable chemical structures at multiple levels relevant to biological interfacing, from fluorescence yield to their influence on cellular membrane potential. We systematically compared the performance of conjugated polymer as cast thin films and as nanoparticles stabilized with amphiphilic polyethylene glycol-poly lactic acid-co-glycolic acid (PEG-PLGA). We assessed in both the dark and under illumination the stability of key optoelectronic properties in various environments, including air and biologically relevant physiological saline solutions. We found that photoreduction of oxygen correlates with nanoparticle and film degradation in physiologically relevant media. Using patch-clamp recordings in cell lines and primary neurons, we identified two broad classes of membrane potential response, which correspond to photosensitizer- and photothermal-mediated effects. Last, we introduced a metric named OED50 (optical energy for 50% depolarization), which conveys the phototoxic potency of a given agent and thereby its operational photo-safety profile.

Sub-millisecond Control of Neuronal Firing by Organic Light-Emitting Diodes.

Optogenetics combines optics and genetics to enable minimally invasive cell-type-specific stimulation in living tissue. For the purposes of bio-implantation, there is a need to develop soft, flexible, transparent and highly biocompatible light sources. Organic semiconducting materials have key advantages over their inorganic counterparts, including low Young's moduli, high strain resistances, and wide color tunability. However, until now it has been unclear whether organic light emitting diodes (OLEDs) are capable of providing sufficient optical power for successful neuronal stimulation, while still remaining within a biologically acceptable temperature range. Here we investigate the use of blue polyfluorene- and orange poly(p-phenylenevinylene)-based OLEDs as stimuli for blue-light-activated Sustained Step Function Opsin (SFFO) and red-light-activated ChrimsonR opsin, respectively. We show that, when biased using high frequency (multi-kHz) drive schemes, the OLEDs permit safe and controlled photostimulation of opsin-expressing neurons and were able to control neuronal firing with high temporal-resolution at operating temperatures lower than previously demonstrated.

Layer 5 Callosal Parvalbumin-Expressing Neurons: A Distinct Functional Group of GABAergic Neurons.

Previous studies have shown that parvalbumin-expressing neurons (CC-Parv neurons) connect the two hemispheres of motor and sensory areas via the corpus callosum, and are a functional part of the cortical circuit. Here we test the hypothesis that layer 5 CC-Parv neurons possess anatomical and molecular mechanisms which dampen excitability and modulate the gating of interhemispheric inhibition. In order to investigate this hypothesis we use viral tracing to determine the anatomical and electrophysiological properties of layer 5 CC-Parv and parvalbumin-expressing (Parv) neurons of the mouse auditory cortex (AC). Here we show that layer 5 CC-Parv neurons had larger dendritic fields characterized by longer dendrites that branched farther from the soma, whereas layer 5 Parv neurons had smaller dendritic fields characterized by shorter dendrites that branched nearer to the soma. The layer 5 CC-Parv neurons are characterized by delayed action potential (AP) responses to threshold currents, lower firing rates, and lower instantaneous frequencies compared to the layer 5 Parv neurons. Kv1.1 containing K+ channels are the main source of the AP repolarization of the layer 5 CC-Parv and have a major role in determining both the spike delayed response, firing rate and instantaneous frequency of these neurons.

Use of SU8 as a stable and biocompatible adhesion layer for gold bioelectrodes.

Gold is the most widely used electrode material for bioelectronic applications due to its high electrical conductivity, good chemical stability and proven biocompatibility. However, it adheres only weakly to widely used substrate materials such as glass and silicon oxide, typically requiring the use of a thin layer of chromium between the substrate and the metal to achieve adequate adhesion. Unfortunately, this approach can reduce biocompatibility relative to pure gold films due to the risk of the underlying layer of chromium becoming exposed. Here we report on an alternative adhesion layer for gold and other metals formed from a thin layer of the negative-tone photoresist SU-8, which we find to be significantly less cytotoxic than chromium, being broadly comparable to bare glass in terms of its biocompatibility. Various treatment protocols for SU-8 were investigated, with a view to attaining high transparency and good mechanical and biochemical stability. Thermal annealing to induce partial cross-linking of the SU-8 film prior to gold deposition, with further annealing after deposition to complete cross-linking, was found to yield the best electrode properties. The optimized glass/SU8-Au electrodes were highly transparent, resilient to delamination, stable in biological culture medium, and exhibited similar biocompatibility to glass.

Nanoparticles: A Challenging Vehicle for Neural Stimulation.

Neurostimulation represents a powerful and well-established tool for the treatment of several diseases affecting the central nervous system. Although, effective in reducing the symptoms or the progression of brain disorders, the poor accessibility of the deepest areas of the brain currently hampers the possibility of a more specific and controlled therapeutic stimulation, depending on invasive surgical approaches and long-term stability, and biocompatibility issues. The massive research of the last decades on nanomaterials and nanoscale devices favored the development of new tools to address the limitations of the available neurostimulation approaches. This mini-review focuses on the employment of nanoparticles for the modulation of the electrophysiological activity of neuronal networks and the related transduction mechanisms underlying the nanostructure-neuron interfaces.

Light-evoked hyperpolarization and silencing of neurons by conjugated polymers.

The ability to control and modulate the action potential firing in neurons represents a powerful tool for neuroscience research and clinical applications. While neuronal excitation has been achieved with many tools, including electrical and optical stimulation, hyperpolarization and neuronal inhibition are typically obtained through patch-clamp or optogenetic manipulations. Here we report the use of conjugated polymer films interfaced with neurons for inducing a light-mediated inhibition of their electrical activity. We show that prolonged illumination of the interface triggers a sustained hyperpolarization of the neuronal membrane that significantly reduces both spontaneous and evoked action potential firing. We demonstrate that the polymeric interface can be activated by either visible or infrared light and is capable of modulating neuronal activity in brain slices and explanted retinas. These findings prove the ability of conjugated polymers to tune neuronal firing and suggest their potential application for the in-vivo modulation of neuronal activity.

Shedding Light on Living Cells.

An overview of the optical methods available to modulate the cellular activity in cell cultures and biological tissues is presented, with a focus on the use of exogenous functional materials that absorb electromagnetic radiation and transduce it into a secondary stimulus for cell excitation, with high temporal and spatial resolution. Both organic and inorganic materials are critically evaluated, for in vitro and in vivo applications. Finally, as a direct practical application of optical-stimulation techniques, the most recent results in the realization of artificial visual implants are discussed.

Photothermal cellular stimulation in functional bio-polymer interfaces.

Hybrid interfaces between organic semiconductors and living tissues represent a new tool for in-vitro and in-vivo applications, bearing a huge potential, from basic researches to clinical applications. In particular, light sensitive conjugated polymers can be exploited as a new approach for optical modulation of cellular activity. In this work we focus on light-induced changes in the membrane potential of Human Embryonic Kidney (HEK-293) cells grown on top of a poly(3-hexylthiophene) (P3HT) thin film. On top of a capacitive charging of the polymer interface, we identify and fully characterize two concomitant mechanisms, leading to membrane depolarization and hyperpolarisation, both mediated by a thermal effect. Our results can be usefully exploited in the creation of a new platform for light-controlled cell manipulation, with possible applications in neuroscience and medicine.