View Section, Volume 1. General Principles. View Section, Introduction to Principles of Toxicology. View Section, 1. View Section, Toxicokinetics. View Section, Mechanisms. View Section, Risk Assessment.
View Section, Index to Volume 1. View Section, Volume 2.
Cellular and Molecular Toxicology. View Section, Basic Principles. View Section, 2. View Section, Receptor Systems. View Section, Alterations in Cell Signaling. View Section, Index to Volume 2. View Section, Volume 3. Toxicology Testing and Evaluation. View Section, 3. View Section, Index to Volume 3. View Section, Volume 4. View Section, 4. View Section, Index to Volume 4. View Section, Volume 5. Immune System Toxicology.
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View Section, 5. View Section, Index to Volume 5. View Section, Volume 6. Cardiovascular Toxicology. View Section, Introduction to Cardiovascular Biology. View Section, 6. View Section, Methods of Analysis. View Section, Index to Volume 6. View Section, Volume 7. Renal Toxicology. View Section, 7. View Section, Index to Volume 7. View Section, Volume 8. Respiratory Toxicology.
View Section, 8. View Section, Index to Volume 8. View Section, Volume 9. Hepatic Toxicology. View Section, 9. View Section, Index to Volume 9. View Section, Volume Gastrointestinal Toxicology. A related but distinct preparation made from single-cell suspensions of neural tissue is the reaggregate culture.
Organic Solvent Neurotoxicity
Instead of being placed in culture dishes and allowed to settle onto the surface of the dishes, the cells are kept in suspension by agitation; under appropriate conditions, they stick to one another and form aggregates of controllable size and composition. Typically, the cells in an aggregate organize themselves and exhibit intercellular relations that are a function of and bear some resemblance to the brain region that was the source of the cells.
The cells establish a three-dimensional, often laminated structure, perhaps approximating the in vivo nervous system more closely than do the dissociated cultures grown on the surface of a dish. Reaggregate cultures lend themselves to large-scale, quantitative experiments in which neurobiologic variables can be examined, although morphologic and ligand-binding studies are performed less readily than with surface cultures.
The Role of Glia in Neurotoxicity, Second Edition - Google Books
Organotypic explant cultures are even more closely related to the intact nervous system. Small pieces or slices of neural tissue are placed in culture and can be maintained for long periods with substantial maintenance of structural and cell-cell relations of intact tissue. Specific synaptic relations develop and can be maintained and evaluated, both morphologically and electro-physiologically. Because all regions of the nervous system are amenable to this sort of preparation, it is possible to analyze toxic agents that are active only in specific regions of the central or peripheral nervous system.
Explants can be made from relatively thin slices of neural tissue, so detailed morphologic and intracellular electrophysiologic studies are possible. Their anatomic integrity is such that they capture many of the cell-cell interactions characteristic of the intact nervous system while allowing a direct, continuing evaluation of the effects of a potentially neurotoxic compound added to the culture medium. The process of myelination has been studied extensively in explant cultures, and considerable neurotoxicologic information has been gained.
As noted above, the pathogenetic actions of excitatory amino acids normally active in the nervous system, as well as such analogues as the neurotoxin BMAA, have been revealed by experiments with organotypic cultures. Organ Cultures. A preparation similar to an explant culture is the organ culture, in which an entire organ, such as the inner ear or a ganglion, rather than slices or frag-. Obviously, only structures so small that their viability is not compromised can be treated in this way.
The advantages of the various types of in vitro systems are summarized in Table Most in vitro preparations are made from young, usually prenatal animals. But cultures derived from human neural tissue have been the object of a number of studies. Typically, a period of rapid change and development occurs immediately after the cultures are initiated, and conditions become much more stable if the cultures are maintained for weeks or months. Thus, the preparations can be used to study neurotoxic effects that might be specific to developing nervous tissue and to compare the effects of agents in developing stable tissue.
In general, the technical ease of maintaining a culture varies inversely with the degree to which it captures a spectrum of in vivo characteristics of nervous system behavior. The problem of biotransformation of potentially neurotoxic compounds is shared by all, although the more complete systems explant or organ cultures might alleviate this problem in specific instances. In many culture systems, complex and ill-defined additives—such as fetal calf serum, horse serum, and human placental serum—are used to promote cell survival. A number of thoroughly described synthetic media are now available, however, and such fully defined culture systems can be used where necessary.
Indicators of neuronal and glial function, and hence indicators of neurotoxicity, are outlined in Table A broad range of in vitro systems are now available for studying development of the nervous system and the normal function of neurons and glial cells. The possible neurotoxic impact of any chemical on any specific neurobiologic variable could, in principle, be screened with an appropriate set of in vitro tests.
In practice, of course, because the number of potential neurobiologic end points to be measured is so large, screening for the effects of any agent on all of them would be prohibitively expensive in time and money. The question, then, is whether a feasible battery of tests will pick up an acceptably large percentage of toxic chemicals while generating an acceptably low percentage of false positives. Ideally, the screen would.
This represents the committees judgment. To what extent could an in vitro system provide such a screening instrument? Two basic questions are associated with the use of in vitro tests for that purpose:. What indicators of neuronal or glial damage would be sufficiently general to be useful? What specific test systems or combinations would be adequate to cover a number of different and differentially site-specific neurotoxic agents?
The first question might be answered by a combination of assays that would include general indicators of neuronal and glial survival and a few more specific indicators. Counts of numbers of surviving neurons or glia and biochemical measurements of tetanus-toxin-specific or sodium-channel-specific ligand binding could be used as general indicators. If a chemical has a single very specific neurologic target, this would in general be missed, but it might be anticipated that such a specific effect would be accompanied by more general secondary neuropathologic consequences.
For instance, if spinal-cord or cerebral cortical cell cultures are exposed to the specific voltage-dependent sodium channel blocker tetrodotoxin for 4—5 days, the decrease in electric activity kills about half the neurons. As to the second question, the choice of culture systems to be used is difficult. It is axiomatic that no cell culture represents a normal nervous system.
Even in cocultured explants, the normal connections among cells are disrupted. Specific neurobiologic properties have been shown not to be expressed in various in vitro preparations where they. For instance, in dissociated hippocampal pyramidal cells, serotonergic responses cannot be demonstrated; the responses appear during development in vivo, but appropriate signals that induce their expression are evidently lacking in vitro.
It is impossible to predict how such departures from normality will influence the screen's ability to detect the effects of a test substance. Some compromise between comprehensiveness and fiscal feasibility would have to be made. Neural and glial cell lines are available and relatively straightforward technically.