Photostimulation

What is photostimulation?
What is FRAP?
What is optogenetics?
Research introduction: Light-controlled tools for intracellular cAMP generation
Highlighted product: LCOS-SLM
Highlighted product: Photomultiplier tube module with gate function
Other imaging techniques

What is photostimulation?

Photostimulation refers to an experimental method where samples such as living organisms or cells are stimulated with light, and the resulting reactions are observed. Various experimental techniques utilize photostimulation, including FRAP (fluorescence recovery after photobleaching), optogenetics, photoconversion, and photoswitching. The following sections describe FRAP and optogenetics.

What is FRAP?

FRAP is the measurement of the time required for the fluorescence brightness of a specific fluorescent molecule in the observation area to recover after photobleaching with strong excitation light. It is a method that can estimate the movement speed of molecules from outside the observation area. The less movement of molecules, the slower the recovery speed of fluorescence, and the more movement of molecules, the faster the recovery speed.

To achieve photobleaching in a specific region, FRAP requires very strong excitation light and optical systems that can precisely target the excitation light to that area. Commonly, lasers used in confocal microscopy are employed for excitation. Spatial light modulators (SLMs) or digital mirror devices (DMDs) are used to focus the laser exclusively on the desired region.

Principle of FRAP

Figure 1: Principle of FRAP

What is optogenetics?

Optogenetics is an interdisciplinary field that combines optics and genetics. It is also known as light genetics. In optogenetics, membrane proteins such as channelrhodopsin and halorhodopsin, which respond to light energy, are expressed in neurons. By irradiating specific wavelengths of light, it is possible to excite or inhibit target neurons. This technique can be applied to elucidate the structure and function of neural networks.

Traditionally, electrical stimulation methods were primarily used to study neural function. However, these methods had a drawback: they stimulated not only the targeted neurons but also neighboring neurons connected to them. Optogenetics overcomes this limitation by allowing precise stimulation of only the neurons expressing membrane proteins like channelrhodopsin. As a result, researchers can observe neural cell movement more accurately.

In optogenetics, there are cases where only specific neurons need to be stimulated with light. In such situations, it is necessary to irradiate light only to specific areas. Additionally, when stimulating multiple neurons simultaneously, light must be directed to multiple areas. Our LCOS-SLM allows customizable illumination patterns, enabling precise light irradiation to specific regions.

Differences between optogenetics and electrical stimulation

Figure 2: Differences between optogenetics and electrical stimulation

Research introduction: Light-controlled tools for intracellular cAMP generation

In recent years, various light manipulation techniques have made significant advancements in the field of life sciences. Since 2006, light-sensitive ion channels have become an essential tool for optogenetics, the manipulation of light, in the field of neuroscience. However, within living organisms, numerous signals exist beyond ion channels. Notably, cyclic nucleotides such as cAMP and cGMP play crucial roles in diverse intracellular signal transduction pathways. In the main signaling pathway involving cAMP, activation of adenylate cyclase or inhibition of phosphodiesterase leads to an increase in intracellular cAMP concentration, subsequently activating protein kinase A (PKA). This activation is followed by the appearance of voltage-dependent Ca²⁺ channels and various cellular responses. Hamamatsu Photonics is researching an optogenetics tool called ‘PAC’ that allows light-controlled cAMP generation to induce these responses.

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Highlighted product: LCOS-SLM

The LCOS-SLM (liquid crystal on silicon - spatial light modulator) is a device that allows electrical control of the phase of laser light. It consists of a structure where a liquid crystal is sandwiched between a CMOS chip with pixel electrodes arranged in a two-dimensional pattern and transparent electrodes deposited on a glass substrate. Digital images output from a PC are converted to analog signals by a dedicated driving circuit and applied with voltage to the pixel electrodes on the CMOS chip. By tilting the liquid crystal molecules through this voltage, the refractive index of the liquid crystal changes, allowing precise control of the phase of light irradiated onto each pixel. The LCOS-SLM, capable of high-precision control of the light phase, finds applications ranging from research purposes such as optical vortex generation to industrial applications like aberration correction in microscopy and fine code marking through multipoint branching.

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Highlighted product: Photomultiplier tube module with gate function

A Photomultiplier tube module with gate function is an optoelectronic multiplier tube module with electrically controllable gate operation. By applying gate operation, it is possible to control the timing of light detection. In experiments involving light stimulation, when irradiating the sample with light for stimulation, in addition to the actual fluorescence signal that you want to measure, another light for stimulation is inadvertently detected. To avoid this, it is possible to apply gate functions during light irradiation for stimulation to prevent the detection of unwanted light.