Measurement of neuronal Ca2+ transients using simultaneous microfluorimetry and electrophysiology. 1988

S A Thayer, and M Sturek, and R J Miller
Department of Pharmacological and Physiological Sciences, University of Chicago, IL 60637.

Fluorescent indicator molecules, such as fura-2, are useful probes for studying the concentrations of ions in single cells. A key feature of these fluorescent dyes is the shift in their excitation spectra upon binding the ion, thus making alternate excitation from two wavelengths desirable. In this report we describe an inexpensive system for making simultaneous electrophysiological and dual excitation fluorescence measurements using equipment much of which is available in a typical biophysical laboratory. In order to synchronize the fluorescence signal with the appropriate excitation wavelength we devised a simple computer program which uses the output of photodiodes placed in the excitation beam to determine which wavelength is illuminating the cell. We also describe the use of a liquid light guide to transmit excitation illumination from the light source to the epifluorescence port of the microscope in order to isolate a perfusion chamber from light, electrical noise and vibration. A sensitive light detection system was assembled using a photomultiplier tube and discriminator that took advantage of sampling single unit events obtained with photon counting rather than the analog of anode current. However, rather than employing a sophisticated and expensive photon counting system, a filter was used to integrate the signal so that an analog output could be presented to a multichannel analog to digital converter commonly used for electrophysiological recordings.(ABSTRACT TRUNCATED AT 250 WORDS)

UI MeSH Term Description Entries
D009475 Neurons, Afferent Neurons which conduct NERVE IMPULSES to the CENTRAL NERVOUS SYSTEM. Afferent Neurons,Afferent Neuron,Neuron, Afferent
D002118 Calcium A basic element found in nearly all tissues. It is a member of the alkaline earth family of metals with the atomic symbol Ca, atomic number 20, and atomic weight 40. Calcium is the most abundant mineral in the body and combines with phosphorus to form calcium phosphate in the bones and teeth. It is essential for the normal functioning of nerves and muscles and plays a role in blood coagulation (as factor IV) and in many enzymatic processes. Coagulation Factor IV,Factor IV,Blood Coagulation Factor IV,Calcium-40,Calcium 40,Factor IV, Coagulation
D003592 Cytophotometry A method for the study of certain organic compounds within cells, in situ, by measuring the light intensities of the selectively stained areas of cytoplasm. The compounds studied and their locations in the cells are made to fluoresce and are observed under a microscope. Microfluorometry,Cytophotometries,Microfluorometries
D004594 Electrophysiology The study of the generation and behavior of electrical charges in living organisms particularly the nervous system and the effects of electricity on living organisms.
D005728 Ganglia, Sympathetic Ganglia of the sympathetic nervous system including the paravertebral and the prevertebral ganglia. Among these are the sympathetic chain ganglia, the superior, middle, and inferior cervical ganglia, and the aorticorenal, celiac, and stellate ganglia. Celiac Ganglia,Sympathetic Ganglia,Celiac Ganglion,Ganglion, Sympathetic,Ganglia, Celiac,Ganglion, Celiac,Sympathetic Ganglion
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D001572 Benzofurans Compounds that contain a BENZENE ring fused to a furan ring. Coumarones,Diphenylbenzofuran
D016257 Fura-2 A fluorescent calcium chelating agent which is used to study intracellular calcium in tissues. Fura 2
D051381 Rats The common name for the genus Rattus. Rattus,Rats, Laboratory,Rats, Norway,Rattus norvegicus,Laboratory Rat,Laboratory Rats,Norway Rat,Norway Rats,Rat,Rat, Laboratory,Rat, Norway,norvegicus, Rattus

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