Faculty Bibliography

Two classes of substances exist within the extracellular space: energetic and informational. Examples of the former are glucose, dissolved oxygen and CO2 while the latter include excitatory amino acids, cathecholamines and opiates. The simple ions Na+ and Cl- are generally associated with energetic processes while extracellular K+ and Ca2+ tend to be informational in function. Local release of an informational substance brings about a concentration gradient that causes the substance to be dispersed in the extracellular space by diffusion. This process is modified relative to a free aqueous medium by the constraints of volume fraction, tortuosity and uptake. Volume fraction is defined simply as the fraction of a brain region that is extracellular. If a given quantity of substance is released into a region with a reduced volume fraction then the substance will reach a higher concentration than it would in a free medium. Tortuosity is related to the increase in the path length of the random walk of a diffusing particle due to the necessity to navigate around cellular obstructions. Tortuosity manifests itself as a decrease in the diffusion coefficient. Uptake represents the movement of a substance from the extracellular space to the intracellular. Since initially a concentration gradient exists in this direction and all membranes have some permeability some concentration-dependent uptake always occurs. In addition there exist specific carrier-mediated uptake processes for some substances such as amino acids or catecholamines. In some regions the dispersal process can be dominated by uptake rather than diffusion. While volume fraction, tortuosity and uptake have all been demonstrated by a technique based on the use of radiolabels and other methods, these classical techniques have limited spatial and temporal resolution. The advent of methods based on micro-injection of substances by iontophoresis or pressure and subsequent detection with ion-selective microelectrodes (ISMs) or voltammetric microsensors (VMs) has opened a new window onto the dynamic local behavior of the extracellular space. In the last decade our laboratory and others have studied the migration of the test substances tetramethylammonium, tetraethylammonium, AsF6- and alpha naphthalene sulfonate, the endogenous ions K+ and Ca2+, the epileptogenic agent penicillin and the neurotransmitter dopamine. These studies have been carried out on the cerebellum and some other regions in a variety of species that include rat, turtle, skate and an intervertebrate, the cuttlefish.(ABSTRACT TRUNCATED AT 400 WORDS)

The diffusion of penicillin was studied in agar gel and the cerebral cortex of the rat using pressure microinjection and ion-selective microelectrodes selective to penicillin. From the agar measurements a free diffusion coefficient for penicillin of 3.52 +/- 0.08 (mean +/- S.E.M.) X 10(-6) cm2.s-1 for 37 degrees C was determined. The tortuosity value in the cortex was 1.62 +/- 0.03 (mean +/- S.E.M.) at the same temperature implying an apparent diffusion coefficient of 1.34 +/- 0.07 (mean +/- S.E.M.) x 10(-6) cm2.s-1. This tortuosity value means that penicillin diffuses in the cortex in a similar manner to other extracellular substances. These diffusion values clarify previous estimates and permit accurate evaluation of epilepsy models based on the application of penicillin

The isolated turtle cerebellum was used as a model system to study effects of depolarizing conditions on interstitial ascorbic acid concentration. The depolarizing stimulus was Leao's spreading depression, which is characterized by transient negative extracellular potentials, high potassium levels (20-60 mM), and local depression of neuronal activity. Interstitial concentrations of ascorbate (200-400 microM) and other electroactive species were monitored voltammetrically, using graphite fiber microelectrodes. Total tissue ascorbate (1,810 nmol/g tissue wet weight) was similar to mammalian levels and was several orders of magnitude higher than catecholamine and indoleamine content. During spreading depression, a large (up to 200 microM) increase in concentration of interstitial electroactive species was monitored. Use of Nafion- and ascorbate oxidase-coated electrodes and uricase confirmed that ascorbate was the only substance detected. Simultaneous monitoring of ascorbate, extracellular potential, and extracellular volume (using tetramethylammonium and ion-selective microelectrodes) indicated that (a) the ascorbate increase began with the decrease in extracellular volume during spreading depression, and (b) much of the increase was the result of extracellular volume decrease. In sucrose-substituted medium, in which volume changes are eliminated, a 50 microM increase in interstitial ascorbate, caused by release from intracellular stores, was also seen. The ascorbate concentration increase was prolonged in sucrose medium, suggesting that an uptake process involving sodium may further regulate interstitial ascorbate concentration

A review of some of the literature on Ca2+ diffusion in free media and a variety of nervous tissues is presented. In the majority of tissue studies the apparent diffusion coefficient of Ca2+ is three to nine times smaller than that in a free aqueous medium. The methodology of using pressure microejection and Ca2+ ion-selective microelectrodes to measure Ca2+ diffusion is discussed. Our ongoing studies of Ca2+ diffusion in the cerebral cortex of the rat, using these methods, also confirm that Ca2+ diffusion is mainly influenced by the tortuosity of the tissue rather than other factors such as binding to extracellular charge sites or uptake

We report in vitro experiments on the source(s) of the magnetic fields produced by the brain. Theoretical arguments suggest that the dominant sources should be dipolar and oriented parallel to the scalp. Using an isolated turtle cerebellum as a model, we find that the fields produced following dorsal stimulation are attributable to current flow perpendicular to the cerebellum surface, suggesting Purkinje cell sources. We also discuss observations of longer lasting fields associated with spreading depression induced in the cerebellum

A superconducting sensor was used to measure the magnetic field evoked from the isolated cerebellum of the turtle by brief electrical stimulations of the dorsal surface. The field was generated by neuronal activities, since its amplitude was reduced when Mn2+ was applied and completely abolished with tetrodotoxin. In normal bathing medium, the field amplitude at a distance of 17 mm was as much as 1 pT, demonstrating that the magnetic technique can be used to remotely monitor neural activities in regions of 10 mm3 or even less