Voltage-sensitive dyes, also known as potentiometric dyes, are dyes which change their spectral

Electromagnetic spectrum

The electromagnetic spectrum is the range of all possible frequencies of electromagnetic radiation. The "electromagnetic spectrum" of an object is the characteristic distribution of electromagnetic radiation emitted or absorbed by that particular object....

 properties in response to voltage

Voltage

Voltage, otherwise known as electrical potential difference or electric tension is the difference in electric potential between two points — or the difference in electric potential energy per unit charge between two points...

 changes. They are able to provide linear measurements of firing activity of single neurons, large neuronal populations or activity of myocytes. Many physiological processes are accompanied by changes in cell membrane potential

Membrane potential

Membrane potential is the difference in electrical potential between the interior and exterior of a biological cell. All animal cells are surrounded by a plasma membrane composed of a lipid bilayer with a variety of types of proteins embedded in it...

 which can be detected with voltage sensitive dyes. Measurements may indicate the site of action potential

Action potential

In physiology, an action potential is a short-lasting event in which the electrical membrane potential of a cell rapidly rises and falls, following a consistent trajectory. Action potentials occur in several types of animal cells, called excitable cells, which include neurons, muscle cells, and...

 origin, and measurements of action potential velocity and direction may be obtained.

Potentiometric dyes are used to monitor the electrical activity inside cell organelles where it is not possible to insert an electrode

Electrode

An electrode is an electrical conductor used to make contact with a nonmetallic part of a circuit...

, such as the mitochondria. This technology is especially powerful for the study of patterns of
activity in complex multicellular preparations. It also makes possible the
measurement of spatial and temporal variations in membrane potential

Membrane potential

Membrane potential is the difference in electrical potential between the interior and exterior of a biological cell. All animal cells are surrounded by a plasma membrane composed of a lipid bilayer with a variety of types of proteins embedded in it...

 along the
surface of single cells. Ideally, the electric potential

Electric potential

In classical electromagnetism, the electric potential at a point within a defined space is equal to the electric potential energy at that location divided by the charge there...

 is determined at very high impedance

Electrical impedance

Electrical impedance, or simply impedance, is the measure of the opposition that an electrical circuit presents to the passage of a current when a voltage is applied. In quantitative terms, it is the complex ratio of the voltage to the current in an alternating current circuit...

, so that virtually zero current

Electric current

Electric current is a flow of electric charge through a medium.This charge is typically carried by moving electrons in a conductor such as wire...

 flow occurs to interfere with cellular reactions.

Types of dyes

Fast dyes: These are amphiphilic membrane staining dyes which usually have a pair of
hydrocarbon chains acting as membrane anchors and a hydrophilic group which aligns the chromophore

Chromophore

A chromophore is the part of a molecule responsible for its color. The color arises when a molecule absorbs certain wavelengths of visible light and transmits or reflects others. The chromophore is a region in the molecule where the energy difference between two different molecular orbitals falls...

perpendicular to the membrane/aqueous interface. The chromophore is believed to undergo a large
electronic charge shift as a result of excitation from the ground to the excited state

Excited state

Excitation is an elevation in energy level above an arbitrary baseline energy state. In physics there is a specific technical definition for energy level which is often associated with an atom being excited to an excited state....

 and this underlies the
putative electrochromic mechanism for the sensitivity of these dyes to membrane potential. This molecule (dye) intercalates among the lipophilic

Lipophilic

Lipophilicity, , refers to the ability of a chemical compound to dissolve in fats, oils, lipids, and non-polar solvents such as hexane or toluene. These non-polar solvents are themselves lipophilic — the axiom that like dissolves like generally holds true...

 part of biological membrane

Biological membrane

A biological membrane or biomembrane is an enclosing or separatingmembrane that acts as a selective barrier, within or around a cell. It consists of a lipid bilayer with embedded proteins that may constitute close to 50% of membrane content...

s. This orientation assures that the excitation induced
charge redistribution will occur parallel to the electric field

Electric field

In physics, an electric field surrounds electrically charged particles and time-varying magnetic fields. The electric field depicts the force exerted on other electrically charged objects by the electrically charged particle the field is surrounding...

 within the membrane. A change in the
voltage across the membrane will therefore cause a spectral shift

Redshift

In physics , redshift happens when light seen coming from an object is proportionally increased in wavelength, or shifted to the red end of the spectrum...

 resulting from a direct interaction
between the field and the ground and excited state dipole moments

Magnetic moment

The magnetic moment of a magnet is a quantity that determines the force that the magnet can exert on electric currents and the torque that a magnetic field will exert on it...

.

Examples

Commonly used voltage sensitive dyes are substituted aminonaphthylethenylpyridinium (ANEP) dyes, such as di-4-ANEPPS, di-8-ANEPPS, and RH237. Depending on their chemical modifications which change their physical properties

Physical property

A physical property is any property that is measurable whose value describes a physical system's state. The changes in the physical properties of a system can be used to describe its transformations ....

 they are used for different experimental procedures. They were first described in 1985 by the research group of Leslie Loew. ANNINE-6plus

ANNINE-6plus

ANNINE-6plus is a water soluble voltage sensitive dye . This compound was developed at the Max Planck Institute for Biochemistry in Germany...

 is the latest voltage sensitive dye with fast response (ns response time

Response time

In technology, response time is the time a system or functional unit takes to react to a given input.- Data processing :In data processing, the response time perceived by the end user is the interval between the instant at which an operator at a terminal enters a request for a response from a...

) and high voltage

High voltage

The term high voltage characterizes electrical circuits in which the voltage used is the cause of particular safety concerns and insulation requirements...

 sensitivity. It has been applied to measure the action potentials of a single t-tubule

T-tubule

A T-tubule is a deep invagination of the sarcolemma, which is the plasma membrane, only found in skeletal and cardiac muscle cells...

 of cardiomyocytes by Guixue Bu et al.

Materials

The core material for imaging brain activity with voltage-sensitive dyes are the dyes themselves. These voltage-sensitive dyes are lipophilic

Lipophilic

Lipophilicity, , refers to the ability of a chemical compound to dissolve in fats, oils, lipids, and non-polar solvents such as hexane or toluene. These non-polar solvents are themselves lipophilic — the axiom that like dissolves like generally holds true...

 and preferably localized in membranes with their hydrophobic tails. They are used in applications involving fluorescence or absorption; they are fast acting and are able to provide linear measurements of changes in membrane potential.

A variety of specialized equipment may be used in conjunction with the dyes, and choices in equipment will vary according to the particularities of a preparation. Essentially, equipment will include specialized microscopes and imaging devices, and may include technical lamps or lasers.

Strengths and weaknesses

Strengths of imaging brain activity with voltage-sensitive dyes include the following abilities:

• Measurement of population signals from many areas may be taken simultaneously, and hundreds of neurons may be recorded from. Such multisite recordings may provide precise information on action potential initiation and propagation (including direction and velocity), and on the entire branching structure of a neuron.
• Measurements of spike activity in a ganglion that is producing behaviour can be taken and may provide information about how the behaviour is producing.
• In certain preparations the pharmacological effects of the dyes may be completely reversed by removing the staining pipette and allowing the neuron 1–2 hours for recovery.
• Dyes may be used to analyze signal integration in terminal dendritic branches. Voltage-sensitive dyes offer the only alternative to genetically encoded voltage sensitive proteins (such as Ci-VSP derived proteins) for doing this.

Weaknesses of imaging brain activity with voltage-sensitive dyes include the following problems:

• Voltage-sensitive dyes may respond very differently from one preparation to another; typically tens of dyes must be tested in order to obtain an optimal signal., imaging parameters, such as excitation wavelength, emission wavelength, exposure time, should also be optimized
• Voltage-sensitive dyes often fail to penetrate through connective tissue
  Connective tissue
  "Connective tissue" is a fibrous tissue. It is one of the four traditional classes of tissues . Connective Tissue is found throughout the body.In fact the whole framework of the skeleton and the different specialized connective tissues from the crown of the head to the toes determine the form of...
  
   or move through intracellular spaces to the region of membrane desired for study. Staining is a serious issue in applications of these dyes. Water soluble dyes, such as ANNINE-6plus
  ANNINE-6plus
  ANNINE-6plus is a water soluble voltage sensitive dye . This compound was developed at the Max Planck Institute for Biochemistry in Germany...
  
  , do not suffer this problem.
• Noise is a problem in all preparations with voltage-sensitive dyes and in certain preparations the signal may be significantly obscured. It can be solved with spatial filter
  Spatial filter
  A spatial filter is an optical device which uses the principles of Fourier optics to alter the structure of a beam of coherent light or other electromagnetic radiation. Spatial filtering is commonly used to "clean up" the output of lasers, removing aberrations in the beam due to imperfect, dirty,...
  
  ing algorithms. A practical algorithm to deal with this signai has been published.

• Cells may be permanently affected by treatments. Lasting pharmacological effects are possible, and the photodynamics of the dyes can be damaging.