Neurofilament Light Regulates Axon Caliber, Synaptic Activity, and Organelle Trafficking in Cultured Human Motor Neurons

Neurofilament light (NFL) is one of the proteins forming multimeric neuron-specific intermediate filaments, neurofilaments, which fill the axonal cytoplasm, establish caliber growth, and provide structural support. Dominant missense mutations and recessive nonsense mutations in the neurofilament light gene (NEFL) are among the causes of Charcot-Marie-Tooth (CMT) neuropathy, which affects the peripheral nerves with the longest axons. We previously demonstrated that a neuropathy-causing homozygous nonsense mutation in NEFL led to the absence of NFL in patient-specific neurons. To understand the disease-causing mechanisms, we investigate here the functional effects of NFL loss in human motor neurons differentiated from induced pluripotent stem cells (iPSC). We used genome editing to generate NEFL knockouts and compared them to patient-specific nonsense mutants and isogenic controls. iPSC lacking NFL differentiated efficiently into motor neurons with normal axon growth and regrowth after mechanical axotomy and contained neurofilaments. Electrophysiological analysis revealed that motor neurons without NFL fired spontaneous and evoked action potentials with similar characteristics as controls. However, we found that, in the absence of NFL, human motor neurons 1) had reduced axonal caliber, 2) the amplitude of miniature excitatory postsynaptic currents (mEPSC) was decreased, 3) neurofilament heavy (NFH) levels were reduced and no compensatory increases in other filament subunits were observed, and 4) the movement of mitochondria and to a lesser extent lysosomes was increased. Our findings elaborate the functional roles of NFL in human motor neurons. NFL is not only a structural protein forming neurofilaments and filling the axonal cytoplasm, but our study supports the role of NFL in the regulation of synaptic transmission and organelle trafficking. To rescue the NFL deficiency in the patient-specific nonsense mutant motor neurons, we used three drugs, amlexanox, ataluren (PTC-124), and gentamicin to induce translational read-through or inhibit nonsense-mediated decay. However, the drugs failed to increase the amount of NFL protein to detectable levels and were toxic to iPSC-derived motor neurons.Peer reviewe

Network adaptation improves temporal representation of naturalistic stimuli in drosophila eye: II Mechanisms

Retinal networks must adapt constantly to best present the ever changing visual world to the brain. Here we test the hypothesis that adaptation is a result of different mechanisms at several synaptic connections within the network. In a companion paper (Part I), we showed that adaptation in the photoreceptors (R1-R6) and large monopolar cells (LMC) of the Drosophila eye improves sensitivity to under-represented signals in seconds by enhancing both the amplitude and frequency distribution of LMCs' voltage responses to repeated naturalistic contrast series. In this paper, we show that such adaptation needs both the light-mediated conductance and feedback-mediated synaptic conductance. A faulty feedforward pathway in histamine receptor mutant flies speeds up the LMC output, mimicking extreme light adaptation. A faulty feedback pathway from L2 LMCs to photoreceptors slows down the LMC output, mimicking dark adaptation. These results underline the importance of network adaptation for efficient coding, and as a mechanism for selectively regulating the size and speed of signals in neurons. We suggest that concert action of many different mechanisms and neural connections are responsible for adaptation to visual stimuli. Further, our results demonstrate the need for detailed circuit reconstructions like that of the Drosophila lamina, to understand how networks process information

Visual Coding in Locust Photoreceptors

Information capture by photoreceptors ultimately limits the quality of visual processing in the brain. Using conventional sharp microelectrodes, we studied how locust photoreceptors encode random (white-noise, WN) and naturalistic (1/f stimuli, NS) light patterns in vivo and how this coding changes with mean illumination and ambient temperature. We also examined the role of their plasma membrane in shaping voltage responses. We found that brightening or warming increase and accelerate voltage responses, but reduce noise, enabling photoreceptors to encode more information. For WN stimuli, this was accompanied by broadening of the linear frequency range. On the contrary, with NS the signaling took place within a constant bandwidth, possibly revealing a ‘preference’ for inputs with 1/f statistics. The faster signaling was caused by acceleration of the elementary phototransduction current - leading to bumps - and their distribution. The membrane linearly translated phototransduction currents into voltage responses without limiting the throughput of these messages. As the bumps reflected fast changes in membrane resistance, the data suggest that their shape is predominantly driven by fast changes in the light-gated conductance. On the other hand, the slower bump latency distribution is likely to represent slower enzymatic intracellular reactions. Furthermore, the Q10s of bump duration and latency distribution depended on light intensity. Altogether, this study suggests that biochemical constraints imposed upon signaling change continuously as locust photoreceptors adapt to environmental light and temperature conditions

A Model for the Detection of Moving Targets in Visual Clutter Inspired by Insect Physiology

We present a computational model for target discrimination based on intracellular recordings from neurons in the fly visual system. Determining how insects detect and track small moving features, often against cluttered moving backgrounds, is an intriguing challenge, both from a physiological and a computational perspective. Previous research has characterized higher-order neurons within the fly brain, known as ‘small target motion detectors’ (STMD), that respond robustly to moving features, even when the velocity of the target is matched to the background (i.e. with no relative motion cues). We recorded from intermediate-order neurons in the fly visual system that are well suited as a component along the target detection pathway. This full-wave rectifying, transient cell (RTC) reveals independent adaptation to luminance changes of opposite signs (suggesting separate ON and OFF channels) and fast adaptive temporal mechanisms, similar to other cell types previously described. From this physiological data we have created a numerical model for target discrimination. This model includes nonlinear filtering based on the fly optics, the photoreceptors, the 1st order interneurons (Large Monopolar Cells), and the newly derived parameters for the RTC. We show that our RTC-based target detection model is well matched to properties described for the STMDs, such as contrast sensitivity, height tuning and velocity tuning. The model output shows that the spatiotemporal profile of small targets is sufficiently rare within natural scene imagery to allow our highly nonlinear ‘matched filter’ to successfully detect most targets from the background. Importantly, this model can explain this type of feature discrimination without the need for relative motion cues

Glutamate, GABA and Acetylcholine Signaling Components in the Lamina of the Drosophila Visual System

Synaptic connections of neurons in the Drosophila lamina, the most peripheral synaptic region of the visual system, have been comprehensively described. Although the lamina has been used extensively as a model for the development and plasticity of synaptic connections, the neurotransmitters in these circuits are still poorly known. Thus, to unravel possible neurotransmitter circuits in the lamina of Drosophila we combined Gal4 driven green fluorescent protein in specific lamina neurons with antisera to γ-aminobutyric acid (GABA), glutamic acid decarboxylase, a GABAB type of receptor, L-glutamate, a vesicular glutamate transporter (vGluT), ionotropic and metabotropic glutamate receptors, choline acetyltransferase and a vesicular acetylcholine transporter. We suggest that acetylcholine may be used as a neurotransmitter in both L4 monopolar neurons and a previously unreported type of wide-field tangential neuron (Cha-Tan). GABA is the likely transmitter of centrifugal neurons C2 and C3 and GABAB receptor immunoreactivity is seen on these neurons as well as the Cha-Tan neurons. Based on an rdl-Gal4 line, the ionotropic GABAA receptor subunit RDL may be expressed by L4 neurons and a type of tangential neuron (rdl-Tan). Strong vGluT immunoreactivity was detected in α-processes of amacrine neurons and possibly in the large monopolar neurons L1 and L2. These neurons also express glutamate-like immunoreactivity. However, antisera to ionotropic and metabotropic glutamate receptors did not produce distinct immunosignals in the lamina. In summary, this paper describes novel features of two distinct types of tangential neurons in the Drosophila lamina and assigns putative neurotransmitters and some receptors to a few identified neuron types