Journal: Pflugers Archiv : European journal of physiology

Article Title: Temporal and spatial summation of laser heat stimuli in cultured nociceptive neurons of the rat.

doi: 10.1007/s00424-022-02728-1

Figure Lengend Snippet: Fig. 1 Experimental setup for investigation of diode laser-induced calcium transients in cultured nociceptive neurons of rats. Laser pulses were applied from above through the cover glasses directly to the neurons that adhere to the back of the cover glasses. Extra- cellular solution was perfused below the neurons through the small space between the cover glass and the bottom of the culture dish. A FURA-2 loaded cells as visualized in the usual XY-plane normally used in calcium imaging experiments (left) as well as after three- dimensional XYZ-reconstruction illustrating the cells hanging upside down from the cover glass (right). Note one mid-size DRG neuron surrounded by several smaller satellite cells. B Sketch of the experi- mental setup; dotted lines indicate magnified inset demonstrating the laser stimulation of hanging cells from above; red line indicates weaker refraction of the invisible NIR laser as compared to the vis- ible pilot laser (grey line). C Photography of the experimental set-up. 1 collimator with glass fiber, 2 micromanipulator, 3 culture dish with cover glass, 4 inlet, 5 outlet, 6 extraction system to keep the surface of the cover glass dry

Article Snippet: The fiber collimator that focused the two laser radiations was moved by a computer-controlled motorized micromanipulator (Sutter MP-285, Sutter Instruments, Novato, CA, USA) in 40-nm steps.

Techniques: Cell Culture, Imaging, Extraction

Journal: BioTechniques

Article Title: Automated single-cell electroporation

doi:

Figure Lengend Snippet: (A) Modified current-to-voltage (I–V) converter circuit. Vi, dc/pulse input voltage; Vo, output voltage; Rc, measurement resistance (100 kΩ); Re, electrode resistance (10–20 MΩ); Rm, membrane resistance; Cm, membrane capacitance; Rcl, cleft resistance; R1 and R2, difference amplifier resistance (100 Ωand 1 kΩ, respectively); R3 and R4, amplifier resistance (10 and 470 Ω, respectively). (B) aSCE experimental setup. aSCE consists of micropipet electrode (ME), micromanipulator, charge-coupled device (CCD) camera, I–V converter, computer, and microscope. (C) Features and options in the graphic-user interface. Vm and Rcr are controllable, and options such as saving data and images, micromanipulator speed, camera exposure time, and retraction length are displayed on the front panel. (D) aSCE protocol. After image capture, a cell position for microelectroporation was selected by image-based cell selection. The ME (attached to computer-controlled motorized micromanipulator) was moved to the selected position (x- and y-axis movement). During approach phases, the electrode was moved incrementally in the z direction until it touched and indented the cell membrane (indenting phase). Approach was stopped when the cleft resistance reached the critical resistance value, Rcl (approximately 0.75 Ω). The electroporation phase was then initiated using preselected membrane voltage (Vm) (square-wave pulse frequency, 200 Hz; duration, 1 s; duty ratio, 10%), which induced membrane pore formation and the transport of molecules into the cell via electrophoresis and diffusion. Upon completion of electroporation, the ME was retracted, and data was saved.

Article Snippet: To accomplish these design goals, we integrated a computer-controlled micromanipulator (MP-285; Sutter Instruments, Novato, CA, USA), a high-performance cooled charge-coupled device (CCD) digital imaging camera (Sensicam-ER; Cooke Corporation, Romulus, MI, USA), an A/D board (National Instruments, Austin, TX, USA) , and a modified current-to-voltage-converting circuit ( ) with an IX71 microscope (Olympus, Lehigh, PA, USA). fig ft0 fig mode=article f1 fig/graphic|fig/alternatives/graphic mode="anchored" m1 Open in a separate window Figure 1 caption a7 caption a8 Components of automated single-cell electroporation (aSCE) (A) Modified current-to-voltage (I–V) converter circuit.

Techniques: Modification, Microscopy, Selection, Electroporation, Electrophoresis, Diffusion-based Assay