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Primary cilia are microtubule-based organelles involved in key signaling and sensing processes in eukaryotic cells. Unlike motile cilia, which have been thoroughly studied, the structure and the composition of primary cilia remain largely unexplored despite their fundamental role in development and homeostasis. They have for long been falsely regarded as simplified versions of motile cilia because they lack distinctive elements such as dynein arms, radial spokes, and central pair complex. However, revealing the detailed molecular composition and 3D structure of primary cilia is necessary in order to understand the mechanisms that govern their functions. Such structural investigations are so far being precluded by the challenging preparation of primary cilia for cryo-electron microscopy. Here, we developed an enabling method for investigating the structure of primary cilia at molecular resolution by cryo-electron tomography. We show that the well-known “9+0” arrangement of microtubule doublets is present only at the base of the primary cilium. A few microns away from the base the ciliary architecture changes into an unstructured bundle of EB1-decorated microtubule singlets and some actin filaments. Our results suggest the existence of a previously unobserved crosstalk between actin filaments and microtubules in the primary cilium. Our work provides unprecedented insights into the molecular structure of primary cilia and a general framework for uncovering their molecular composition and function in health and disease. This opens up new possibilities to study aspects of this important organelle that have so far been out of reach.

Primary cilia are microtubule-based organelles that are important for signaling and sensing in eukaryotic cells. Unlike the thoroughly studied motile cilia, the three-dimensional architecture and molecular composition of primary cilia are largely unexplored. Yet, studying these aspects is necessary to understand how primary cilia function in health and disease. We developed an enabling method for investigating the structure of primary cilia isolated from MDCK-II cells at molecular resolution by cryo-electron tomography. We show that the textbook ‘9 + 0’ arrangement of microtubule doublets is only present at the primary cilium base. A few microns out, the architecture changes into an unstructured bundle of EB1-decorated microtubules and actin filaments, putting an end to a long debate on the presence or absence of actin filaments in primary cilia. Our work provides a plethora of insights into the molecular structure of primary cilia and offers a methodological framework to study these important organelles. Primary cilia are isolated from monolayers of mammalian cells in culture for cryo-ET analyses, revealing new features as well as how they differ from motile cilia.

Discharge of a capacitive storage on a load with unipolar pulse is employed for pumping of powerful lasers and also for feeding of electromagnetic launchers, and pulsed high magnetic field facilities. Spark gaps are often used to commute energy on a load. Triggered spark gaps with electrodynamical acceleration of spark channel were developed and investigated by Kovalchuk and colleagues for the unipolar pulse mode. In some applications oscillatory regime (underdamped sinusoidal) has to be realized for the capacitor bank discharge. In particular, it is valid for a pulsed electromagnetic forming (EMF) technology, which is used for assembling welding, cutting, and forming the details of products in various branches of industry. Spark gaps, developed for unipolar discharge, cannot directly be employed in under-damped (oscillatory) regime, because at current transition through zero the arc channel could stop motion and ignite at initial place on the following half period. Process of the arc ignition and motion became more complicated in the oscillatory regime of capacitor bank discharge. Compact gas switch, intended for operation in oscillatory (low damping) regime of discharge, was introduced later. It is two-electrode switch with electrodynamic acceleration of a spark channel and a matched series injection trigger generator. Two operations regimes have been investigated, namely “fast” regime with current amplitude ~ 160 KA, total charge~ 12 C, period of oscillations 60 µS, full pulse length ~ 400 µs and “slow” regime with current amplitude ~ 30 KA, total charge 18 C, period of oscillations 360 µS, full pulse length ~ 3 ms. The spark gap is designed for 50 kV charging voltage, at a current up to 200 kA, and up to 20 C charge transfer. Two main objectives were pursued in this work: 1. develop test bed for investigation of the spark channel motion for the oscillatory regime of capacitor bank discharge; 2. Investigate arc motion depending on the current and charge transfer in wide operation range and compare it with simulations. In this report we present the test bed schematics and results of the tests.