Jay Boyd Best

Electron microscopic studies of planarian regeneration. I. Fine structure of neoblasts in Dugesia dorotocephala.

Planarian neoblasts were studied during the course of regeneration by electron microscopy. The greatest number of neoblasts were found not in the new blastema of regeneration but in the region of old tissue near the wound surface after about 45–55 hours after decapitation. At around this time they tended to occur in small clusters in this region. The immature neoblast, in accord with previous light microscope descriptions, was found to have a large nucleus, and scanty cytoplasm with numerous free ribosomes. Contrary to previous light microscope descriptions, they contain mitochondria of conventional size and appearance but these are distributed close to the nuclear membrane. They contain little or no endoplasmic reticulum. Differentiating neoblasts show polysomal aggregations of ribosomes, prominent supernumerary nucleoli, endoplasmic reticulum forming at the nuclear membrane, and a curious kind of dense lump in close proximity to the nuclear membrane on the cytoplasmic side. These dense lumps tend to be closely associated with several mitochondria and nestled into a shallow invagination of the nuclear membrane. Fibrous strands connecting the dense lump to adjacent nuclear membrane pores were observed. Annulate lamellae were observed in the differentiating neoblast. The significance of these structural components of the neoblast is discussed.

Electron microscopic studies on planaria: II. Fine structure of the neurosecretory system in the planarian Dugesia dorotocephala

The neurosecretory system in the head of the planarian Dugesia dorotocephala was studied by electron microscopy. Most of the nerve fibers in the neuropil of the brain are unmyelinated and contain organelles such as neurofilaments, synaptic vesicles, and mitochondria. Some of these contain round, electron dense granules (400–1100 A in diameter), with a limiting membrane, that are apparently neurosecretory granules. These were also found in a number of the end bulbs as well as in the perikaryon of the soma of some of the neurons. There is considerable evidence that these are formed in the soma of the neurosecretory cells and migrate out along the nerve fibers to the terminal bulbs. The neurosecretory granules seem to be organized by the Golgi complex, the membrane with a portion of its dense material being formed by vesiculation and budding of the Golgi tubes with concentration of dense material inside the membrane continuing after its formation by vesiculation. The secretory neuron appears to pass from one morphological phase to another in a manner related to the formation and dissolution of Golgi complex and endoplasmic reticulum and the production of neurosecretory granules. These processes seem isomorphic with those associated with neurosecretory activity in higher animals.

Fissioning in Planarians: Control by the Brain

Reduced population densities lead to increased rates of fissioning in planarians whereas higher population densities suppress fissioning. This effect is not primarily due to mucus deposition or substances secreted into the water. Experiments are presented which show a system of population feedback control. In the presence of other planarians, the brain exerts an influence (probably neurohormonal) to suppress fissioning. This influence becomes attenuated with axial distance from the brain.

Toxicology of planarians

Potential advantages of using planarians such as Dugesia dorotocephala (Woodworth) to assay a number of different kinds of toxic effects are illustrated by our experiments on neurotoxicity of nicotine, developmental neurotoxicity and behavioral teratogenesis of methylmercuric chloride (MMC), and carcinogenesis of Cd and tetradecanoyl-phorbol-acetate (TPA). Various concentrations of nicotine in the culture water in which planarians were maintained produced acute neurotoxic effects, evident as disturbances in locomotor and fissioning behavior; planarians also acquired tolerance to nicotine. Behavioral teratogenesis in planarians is illustrated by the effect of subteratogenic concentrations of MMC on regeneration of the brain and behavior of decapitated specimens. Despite normal appearance of heads and brains of MMC-treated regenerates, significant deficits were noted in righting response, locomotion, and prey-catching behavior. EM observations revealed quantitative deficits in brain synapses of MMC-treated regenerates. Exposure to the carcinogens Cd and TPA produced a single kind of neoplastic tumor, a malignant reticuloma, in 76% of specimens treated jointly with sublethal concentrations of both chemicals. Toxicological responses of planarians can be compared to those of vertebrates.

Complex synaptic configurations in planarian brain.

Complex synaptic configurations which appear to have especial evolutionary and functional significance are shown in the neuropil of the brain of the planarian Dugesia dorotocephala. Some of the endings in these synaptic attachments contain dense core vesicles, suggesting that nonadrenaline or serotonin or both are neurotransmitters at a more primitive phyletic level than reported hitherto. The spatial proximity and connectivity of the synapses suggest modes of action permitting greater functional complexity to the planarian brain than previously supposed. Closely adjacent cellular processes which contain polysomal ribosomes, unusual in the neuropil, suggest synaptic transmission-protein synthesis coupling and a possible role in memory.

Fine structure and function of planarian goblet cells

Planarians secrete quantities of mucus that appears to contain one or more glycoproteins as its major constituent. The fine structure associated with the various maturational and functional stages of a type of secretory cell of the planarian Dugesia dorotocephala is described which appears to be isomorphic with the goblet cells involved in the mucous secretion of mammals. This type of secretory cell, which occurs in large numbers in D. dorotocephala , differs from other cells which have been implicated in planarian mucous secretion. It originates and matures in the mesenchymal regions of the planarian, then migrates through the ectodermal basement membrane into the ducts penetrating the ectodermal cell layer. In its early stages it has a pronounced laminar endoplasmic reticulum and extensive Golgi complex activity. Electron transparent vesicles bud off from the tubes of the Golgi complex. These vesicles then, by swelling or coalescing, form the large mucus-filled Golgi vacuoles which almost completely fill the cytoplasm of the mature cell.

Acute toxic responses of the freshwater planarian,Dugesia dorotocephala, to methylmercury

Toxic responses of planaria to various aquatic habitat concentrations of methylmercury chloride (MMC) were investigated. One hundred percent lethality occurred within 5 h in 2 ppM MMC, 24 h in 1 ppM MMC, and 5 days in 0.5 ppM MMC. No deaths occurred in 0.2 ppM MMC over a 10 day period, however, non-lethal toxic responses were observed. Varying degrees of head resorption, progressing caudally from the snout were observed. With continuing exposure, partial head regeneration and recovery toward more normal appearance occurred by 10 days. Teratogenic effects were observed in surgical decapitation experiments. Head regeneration was retarded in 0.1 and 0.2 ppM MMC. Malformations, visible lesions, or gross behavioral abnormalities were produced by 2 week exposure of planaria to concentrations of 20 ppB MMC or lower. (RJC)

Cephalic mechanism for social control of fissioning in planarians: III. Central nervous system centers of facilitation and inhibition.

Previous studies have indicated that asexual reproduction (fissioning) in the planarian Dugesia dorotocephala is socially controlled through a cephalic mechanism: Isolation releases fissioning; grouping inhibits it; decapitation, at the level of the auricles, releases it even in grouped subjects. The brain is not necessary for programming the actual events of fissioning; these are orchestrated by the segmental plexus fissioning (SPF) system. Various surgical cuts were made to ablate selected portions of the central nervous system of isolated and grouped planarians in order to ascertain the inhibitory or facilitatory effects of these in the physiological mediation of such control on the SPF system. These results were synthesized into a model of this control system; the anterior lobes and optic regions of the brain inhibit the SPF system, and the anterior and caudal segmental plexuses facilitate it. These influences are partially tonic and partially contingent upon social stimulation.

Transphyletic Animal Similarities and Predictive Toxicology

Several years ago, I published an article entitled “The Evolution and Organization of Sentient Biological Behavior Systems” in one of the books edited by Wolfgang Yourgrau.1 In that article, I presented the rudiments of a theory to account for the “higher behaviors” that I had observed in the primitive flatworm, planaria. Although a variety of subsequent evidence, in addition to that which was available to me at the time, has lent further support for it and I am even more convinced now than then of its essential correctness, it was, I realize with hindsight, a pretty wild theory relative to the prevailing views of the time. Wolfgang exhibited a great deal of courage in acting as editorial midwife for its delivery. In his other role as “pediatrician” of ideas, I hope that he would not be displeased with this sequel describing the further growth of “the child.”

The Neuroanatomy of the Planarian Brain and Some Functional Implications

The planarians have been used extensively for studies on regeneration, morphogenesis, and, in the last decade, for provocative experiments on the chemical and cellular bases of memory. Yet, in spite of this interest in certain selected classes of phenomena which the planarians manifest, the anatomical and physiological substratum underlying these has been only imperfectly studied or known. This gap in the critical information needed to decipher fully this, now reasonably numerous, body of experiments has rendered it less valuable than it has the potential to be.