Overlap of concanavalin-A and insulin imprinting in rat thymocytes

The thymocytic insulin binding in rats treated with the hormone neonatally on a single occasion increased considerably compared to the untreated control by 3 months of age. Treatment with Concanavalin-A also accounted for an increase in adult insulin binding, whereas neonatal treatment with insulin did not alter the binding relations of Concanavalin-A in adulthood.     

Receptor amplifying effect of serotonin and serotonin analogues in a protozoan (Tetrahymena) model system

A 72-hour treatment of Tetrahymena with serotonin analogues at a concentration of 10(-9) mmole/l resulted in a decrease of the reproduction rate, whereas serotonin itself was ineffective. On the second exposure to antagonists, cellular division was enhanced but its rate remained around or below the control value, while the second exposure to serotonin produced a more marked effect than the first one did, which finding implies a receptor amplification. The most pronounced receptor amplification was caused by the first exposure to serotonin. This observation indicates that the receptor is rather selective as regards the amplification effect.     

Effect of the cAMP level on hormonal imprinting in Tetrahymena

Augmentation of the cAMP level has no positive effect on hormonal imprinting in Tetrahymena. Artificial elevation of the cAMP level may inhibit the development of imprinting or may result in abnormal imprinting. The role of Ca2+ is of great importance in mediation of the imprinting mechanism. Generally, this role is not influenced by an elevated cAMP level but, exceptionally, the latter may effect the mechanism of imprinting.     

Effect of vanadate and ouabain on insulin binding and insulin imprinting in Tetrahymena.

Na-metavanadate and ouabain that act on Na+K(+)-ATPase had no influence on insulin binding to Tetrahymena immediately after treatment, but after 24 h considerably enhanced the binding capacity of generations of progeny. The increase in binding was of a similar magnitude to that elicited by insulin imprinting. Vanadate failed to increase the imprinting potential of insulin while ouabain even prevented insulin imprinting when administered together with insulin, but, did not affect imprinting when administered after insulin. By analogy with higher organisms it appears that inhibition of Na+K(+)-ATPase plays no role in the insulin-like effect of vanadate on the unicellular Tetrahymena, as judged also from the capacity to bind insulin of the generations of offspring.     

Taxon-dependence of receptor level cell-to-cell communication in Tetrahymena: possible explanation for the transmission of hormonal imprinting

Insulin treatment induced in Tetrahymena pyriformis a positive hormonal imprinting, and in Tetrahymena thermophila a negative imprinting, resulting in increased and decreased binding capacity, respectively, at re-exposure to the hormone. The imprinting, or the information associated with it, is transferred by the nutrient medium of the insulin-treated cells to those not treated. The issue of transfer depends on the nature of the receiver taxon, leading always to a positive imprinting in Tetrahymena pyriformis, and to a negative imprinting in Tetrahymena thermophila, regardless of the nature of the 'imprinted' transmitter taxon. The findings substantiate the transferability of hormonal imprinting by the nutrient medium at the unicellular level, the key role of the postreceptorial mechanism in determining the trend of imprinting and may explain the persistence of imprinting in the progeny generations.     

Insulin treatment (hormonal imprinting) increases the insulin production of the unicellular Tetrahymena long term. Is there a simultaneous formation of hormone receptor and hormone?

Insulin treatment of Tetrahymena pyriformis resulted in a long-lasting increase in the insulin content of the cells. After about 200 generations, insulin levels significantly higher than in the control were demonstrated by using a quantitative immunocytochemical method. Although the insulin content fluctuated from day to day, it was always higher in the insulin pre-treated (imprinted) cells than in the controls. The results emphasise the simultaneous stimulation of hormone and receptor formation by hormonal imprinting.     

Effect of insulin imprinting on the 3H-amino acid uptake of the Tetrahymena

Insulin imprinting given to the unicellular Tetrahymena considerably increases the uptake and intracellular storage of amino acids even many generations after the actual contact with the hormone. On the other hand, both the first and the second contacts with insulin increase the rate of the excretion of the stored amino acids. On the basis of the results obtained it seems to be possible that both protein synthesis and exocytosis of the Tetrahymena change as an effect of imprinting, either in general or specifically due to the formation of new hormone receptors.     

Influence of hormone concentration and time factor on development of receptor memory in a unicellular (Tetrahymena) model system

1. Treatment of Tetrahymena pyriformis cells with diiodotyrosine (T2) gave rise to a considerable, concentration-dependent increase of the growth rate within the range of 10(-15) and 10(-9) M, but did not influence it at the level of 10(-18) M. 2. Re-exposure of the cells 1, 2 and 4 weeks later to the hormone concentrations originally used accounted for a marked increase of growth rate at all hormone levels tested, indicating that the extremely low concentration of 10(-18) M, which failed to stimulate growth on first exposure, did nevertheless give rise to hormonal imprinting, which caused the cells to "remember" the hormone, as judged from their increased responsiveness to it on re-exposure. 3. The degree of growth response was concentration-dependent on both first and second exposure: higher levels of treatment gave rise to firmer imprinting, and to greater response on re-exposure. 4. The length of exposure time proved to be more decisive than the level of treatment in respect of the development of hormonal imprinting. 5. Short-term exposures up to 60 min, although they stimulated cell growth by direct effect, gave rise to lasting inhibition of cellular response to re-exposure(s) rather than to hormonal imprinting.     

Effect of inhibitors and activators of tyrosine kinase on insulin imprinting in Tetrahymena.

Primary exposure of Tetrahymena cells to insulin gave rise to hormonal (insulin) imprinting in the offspring generations, as judged from the increase in binding upon reexposure to insulin. Vanadate mimicked the action of insulin, inasmuch as it also induced imprinting for insulin, whereas the other tyrosine kinase activator tested, namely H2O2, had no such effect. However, combined treatment with vanadate+H2O2 + insulin induced a more pronounced imprinting for insulin than either insulin or vanadate on their own. The tyrosine kinase inhibitor genistein, a plant flavonoid, did not change the value for insulin binding significantly relative to the control immediately after exposure, but increased it slightly in the offspring generations after 24 h at high dilution. Upon combination with insulin, 10(-4)M genistein inhibited imprinting by insulin. These experimental observations suggest that there may be a key role for tyrosine kinase activity in the mechanism (development) of imprinting.     

Analysis of expressed sequence tags (ESTs) in the ciliated protozoan Tetrahymena thermophila

To assess the utility of expressed sequence tag (EST) sequencing as a method of gene discovery in the ciliated protozoan Tetrahymena thermophila, we have sequenced either the 5' or 3' ends of 157 clones chosen at random from two cDNA libraries constructed from the mRNA of vegetatively growing cultures. Of 116 total non-redundant clones, 8.6% represented genes previously cloned in Tetrahymena. Fifty-two percent had significant identity to genes from other organisms represented in GenBank, of which 92% matched human proteins. Intriguing matches include an opioid-regulated protein, a glutamate-binding protein for an NMDA-receptor, and a stem-cell maintenance protein. Eleven-percent of the non-Tetrahymena specific matches were to genes present in humans and other mammals but not found in other model unicellular eukaryotes, including the completely sequenced Saccharomyces cerevisiae. Our data reinforce the fact that Tetrahymena is an excellent unicellular model system for studying many aspects of animal biology and is poised to become an important model system for genome-scale gene discovery and functional analysis.     

Impact of a single insulin treatment (imprinting) applied during liver regeneration on hepatic insulin receptor development, blood glucose level and liver function parameters in adult rats.

Rats subjected to partial hepatectomy (surgical removal of two thirds of the liver) showed no appreciable change in serum cholesterol, bilirubin, albumin, total protein and A/G values at 2, 5, 12 and 21 days after the intervention. The enzyme activities characteristic of liver damage (GOT, GPT, LDH, AP) were high in the control group and low in the insulin-imprinted group at 2 days, tended to normalize in both groups at 5 days and changed slightly at 12 days. The blood glucose level was markedly decreased in the control group and to a lesser degree also in the experimental group at 2 and 5 days of sampling. Insulin treatment (loading) performed at 2 and 5 days accounted for a drop of blood glucose which was followed by normalization within 2 h. Starving value and response to insulin loading uniformly fell into the physiological range at 21 days, whereas at 12 days no normalization occurred in either group within 2 h of insulin loading, although the starving value was physiological. The binding capacity of the insulin receptor was markedly low in the control group as long as 12 days, and tended to normalize by 21 days. In the insulin-imprinted group the binding capacity increased over the control at 2 and 5 days and normalized by 12 days.     

Digoxin effect and imprinting in the unicellular Tetrahymena

Treatment with digoxin increased, whereas treatment with ouabain decreased, the growth rate of Tetrahymena, but the progeny generations showed an increased mitotic rate after both treatments. A single exposure to digoxin or ouabain accounted for a more than twofold increase in the incorporation (binding) of the cardiac glycosides in the progeny generations.     

Hormonal imprinting in cell culture II. Evidence of hormonal imprinting and thyrotropin (TSH) -gonadotropin (FSH) overlap in a Chinese hamster ovary (CHO) cell line

Reexposure of cultures of the Chinese hamster ovarian cell line CHO K1 to FITC-labeled hormone 48 h after the first 24-h exposure to FSH or TSH showed that hormonal imprinting, accounting for a greater binding capacity on reexposure, also took place in in vitro conditions. TSH amplified the receptors of FSH to a greater degree than FSH itself, although the reverse effect failed to happen. TSH was able to bind the ovarian cells at first exposure, and to amplify the receptors for itself and--remarkably--to a considerably greater degree for FSH, exactly as observed earlier in in vivo systems.     

Insulin pretreatment (imprinting) produces elevated capacity in the insulin binding of Tetrahymena. Different binding by the cilia of the body and oral field

Gold-labeled insulin is bound first of all to the cilia of the oral field of Tetrahymena. A primary treatment (hormonal imprinting) with insulin increases the binding capacity even after 24h and makes it more sensitive for appearance a week later, within a minute of giving insulin-gold. The food vacuoles contain insulin-gold in pretreated cells or without pretreatment as well, though in imprinted situations the label can be found in pinocytotic vesicles at the bases of cilia in the oral field. Altogether, a functional difference can be observed between the cilia of the oral and non-oral surfaces of Tetrahymena and hormonal imprinting has a specifying effect on the binding of labeled hormone.