Irradiated Explants Research Articles

Acute gamma irradiated Stevia rebaudiana Bertoni enhanced particular types of steviol glycosides

Stevia rebaudiana Bertoni from Asteraceae family is commercially valuable for its steviol glycosides (SGs) contents, which is 300 times sweeter than commercial sugar. The bottleneck in Malaysia is the lack of suitable stevia varieties that are able to thrive well under her climatic conditions and still produce high SGs. Mutation induction including gamma irradiation is effective in generating genetic variations and developing new plant varieties with desired traits. This study was aimed to determine the effects of acute gamma irradiation on phenotypic changes and enhancement of SGs contents of Stevia rebaudiana Bertoni variety AKH L1 (herein after will be designated as AKH L1). In vitro shoot tip explants of AKH L1 were subjected to a gamma doses regime of 10Gy to 50Gy, following which phenotypic changes of the irradiated explants and subsequent regenerated plantlets were observed. All irradiated explants exhibited different survival rates, with the lowest at 9.33±8.33% when subjected to 50Gy, while all the control (non-irradiated explants) survived. The LD50 was found to be at 23Gy. Subsequent irradiation of 900 shoot tip explants at 23Gy, produced 468 surviving shoot tips, which were all capable to develop and successfully sub-cultured until the fourth generation, M4. These M4 in vitro mutant plantlets exhibited significant increase in the numbers of leaf (16.07±5.19) and average leaf size (1.12±0.26cm x 0.54±0.15cm). HPLC analysis performed in parallel further revealed the mutant plants contained higher concentrations of stevioside (387.04ppm), rebaudioside A (670.18ppm) and rebaudioside D (106.26ppm) compared to the non-irradiated plantlets, which exhibited 96.87, 194.42 and 28.25ppm, respectively.

Efecto de diferentes explantes irradiados en la regeneración <i>in vitro</i> de frijol común <i>(Phaseolus vulgaris </i>L.) cultivar “ICA Pijao”

El mejoramiento genético convencional en frijol común resulta difícil debido a que presenta una base genética estrecha y muy estable. En este sentido, la combinación de la mutagénesis y el cultivo de tejidos, es una alternativa para inducir variabilidad genética en la búsqueda de tolerancia a factores bióticos y abióticos. Es por ello, que el presente trabajo tuvo como objetivo determinar el efecto de diferentes explantes irradiados en la regeneración in vitro de frijol común (Phaseolus vulgaris L.) cultivar “ICA Pijao”. Se aplicaron radiaciones gamma en callos, en el nudo cotiledonal con un cotiledón (NC-1) con dosis de 0, 10, 20, 30, 40, 50 y 60 Gy y semillas con 0, 100, 200, 300 y 400 Gy. Se evaluó el porcentaje de germinación, longitud de las raíces, porcentaje de explantes que formaron callos, masa fresca (g) de los callos, número de brotes por callo y el número de brotes con raíces. La radiación gamma provocó una disminución en la masa fresca del callo y NC-1. Los callos y el NC-1 solamente formaron brotes con las dosis de 10 y 20 Gy, pero estos fueron hiperhíricos. Los resultados demostraron que la semilla irradiada fue el explante con el que se logró la regeneración in vitro de plantas con hojas definidas, por lo que se recomienda como explante inicial para el uso combinado de mutagénesis y regeneración in vitro de plantas para el cultivar P. vulgaris “ICA Pijao” a través de la organogénesis indirecta.

Selection of mutants obtained by gamma ray irradiation and analysis of genetic variation using RAPD markers in Acorus calamus L.

Acorus calamus is a perennial and medicinal hydrophyte belonging to the family Acoraceae. Irradiation of gamma ray was carried out to obtain mutants of A. calamus due to their limited genetic variation. In vitro explants of A. calamus were irradiated with various doses (20–500 Gy) of gamma rays and the irradiated explants were subcultured for three cycles (vM3). All these explants were cultured on MS medium (salts and vitamins) with 3% (w/v) sucrose, 0.8% (w/v) agar and 4 mg∙L−1 BA (6-benzyl amino purine). This system was performed to select various mutants such as dwarf and salt toleranct mutants. Four solid dwarf mutants were obtained from 3,844 explants and three salt tolerant mutants were selected among 2,800 explants. In selected mutants, polymorphic bands were obtained by using RAPD analysis with selected 9 random primers. The dwarf and salt tolerant mutants were differentiated as a result of cluster analysis. We obtained four dwarf and three salt tolerant mutants from gamma ray irradiation. We conclude that gamma ray irradiation applied to in vitro culture system is an effective way for inducing exclusive mutantationsof A. calamus and the mutants were cleanly differentiated from the wild-type control through RAPD analysis.

Effects of low dose X‐ray irradiation on porcine articular cartilage explants

Ionizing radiation therapy is a crucial treatment for cancer, but can damage surrounding normal tissues. Damage to articular cartilage leading to arthropathy can occur at irradiated sites. It is unclear whether this response is due to damaging surrounding skeletal structures or direct effects on cartilage. In this study, we showed that irradiation with 2 Gy of X-rays causes a significant reduction in the stiffness of porcine explants 1 week post-irradiation. By using both microindentation and indentation-type atomic force microscopy, ionizing radiation reduces stiffness in both the superficial zone, and throughout the entire thickness of the tissue. Young's modulus values were 75% and 60% lower in 2 Gy irradiated samples when compared with controls using microindentation and nanoindentation, respectively. Glycosaminoglycans (GAGs) released into the culture media of irradiated samples was nearly 100% greater at 24 h after exposure. While collagen content in the tissue is similar between groups, GAG content is 55% lower in irradiated explants compared with controls 7 days after exposure. Therefore, the irradiated explants are unable to recover from the initial loss of GAGs by 1 week. This acute loss of GAGs is a likely contributor to the reduction in modulus seen after exposure to ionizing radiation.

PENGARUH RADIASI SINAR GAMMA TERHADAP KULTUR IN VITRO TANAMAN JAHE

In vitro shoot explants of white ginger and red ginger are irradiated by two different techniques. The first, the tuber explants are irradiated by 2 levels of gamma rays namely 10 and 30 Gy. The second, the shoot explants are irradiated by 2 levels of gamma rays namely 7,5 and 12,5 Gy. The irradiated explants are regenerated on modified MS medium with BAP 2 ppm and NAA 0,25 ppm. The result show that the irradiated tuber explants with 10 and 30 Gy could not initiate new regenerant of ginger. On the other hand the irradiated shoot explants with 7,5 and 12,5 Gy have been able to influence the growth and development of shoot, leaf and root. It is indicated that the level of gamma rays irradiation on shoot explants influences the induced putative mutant of ginger. However, the sprouting of red ginger on 12,5 Gy is higher and faster than 7,5 Gy, and white ginger as well as control. The other morphological characters have not been able to be identified, the experiment are still in process to detect the mutagenic influences.

Towards in vitro selection studies for salinity tolerance in Canino apricot cultivar. Effect of gamma irradiation on in vitro mutation and selection for salt-tolerance

In vitro mutation method was used to obtain salt-tolerant clone in apricot. Small propagules of Canino apricot cultivar were irradiated with gamma ray at doses of 0, 10, 25, 35, 50, 75 and 100 Gy. After 30 days from treatment, both the radio sensitivity and post-irradiation recovery were assessed as the number of proliferated shoots per explants, fresh weight of cultures, shoot length and productivity of irradiated explants. A sudden and sharp decrease in the survival percentage occurred with the dose 75 Gy, while the highest dose (100 Gy) was lethal for all propagules. A marked decline in the number of regenerated shoots per explant and fresh weight of produced cultures was associated with an increase of irradiation doses. Doses in the range of 10-75 Gy, which preserved high survival percentage of irradiated explants, seemed to be more suitable for in vitro mutation in Canino apricot cultivar. Irradiated shoots were exposed to different concentrations of NaCl which were added to the multiplication medium at the rates of 25, 50, 75, 100, 125 mM and after 30 days, vigorous shoots were selected from salinity treatments. In conclusion, apricot tissues exposed to different doses of gamma irradiation in the range of 10-75 Gy, followed by culturing the plantlets produced in a medium containing additional salts (ranging from 25 to 125 mM) can be considered a good method to identify the most tolerant mutants to salts in apricot cultivars.

Caspase‐3 Activation and DNA Damage in Pig Skin Organ Culture After Solar Irradiation

In the present study, a convenient and easy-to-handle skin organ culture was developed from domestic pig ears using polycarbonate Transwell culture inserts in 12-well plate. This alternative model was then tested for its suitability in analyzing the short-term effects of a single solar radiation dose (from 55 to 275 kJ.m(-2)). Differentiation of the pig skin was maintained for up to 48 h in culture, and its morphology was similar to that of fresh human skin. Solar irradiation induced a significant release of the cytosolic enzymes lactate dehydrogenase and extracellular signal-related kinase 2 protein in the culture medium 24 h after exposure. These photocytotoxic effects were associated with the formation of sunburn cells, thymine dimers and DNA strand breaks in both the epidermis and dermis. Interestingly, cell death was dose dependent and associated with p53 protein upregulation and strong caspase-3 activation in the basal epidermis. None of these cellular responses was observed in non-irradiated skin. Finally, topical application of a broad-spectrum UVB + A sunfilter formulation afforded efficient photoprotection in irradiated explants. Thus, the ex vivo pig ear skin culture may be a useful tool in the assessment of solar radiation-induced DNA damage and apoptosis, and for evaluating the efficacy of sunscreen formulations.

Effect of radiation on regeneration of Chinese narcissus and analysis of genetic variation with AFLP and RAPD markers

The aim of our study was to establish an efficient in vitro propagation protocol for Chinese narcissus (Narcissus tazetta var. chinensis) to obtain variants of this species using γ-radiation treatment and evaluate the effectiveness of this system for variant induction using amplification fragment length polymorphism (AFLP) and randomly amplified polymorphic DNA (RAPD) analysis. Various doses (5–100 Gy) of gamma rays were applied to investigate the effect of radiation on adventitious bud formation from bulb-scales and the survival rate of plantlets. It was demonstrated that the regeneration of Chinese narcissus was very sensitive to gamma radiation even at low doses. The survival and multiplication rate significantly decreased with an increase of radiation dose. The optimal irradiation dose for survival and mutation induction was approximately 10 Gy. The genetic variations among the regenerants derived from irradiated explants were evaluated by DNA fingerprinting using RAPD and AFLP markers which detected a variation frequency of 8.33% and 15.48% respectively. The high frequency of mutants detected by molecular markers indicated that treatment of in vitro cultures with γ-rays may be an effective way to improve narcissus cultivars.

Further Investigation of the Response of Human Uroepithelium to Low Doses of Cobalt-60 Gamma Radiation

Explant cultures of normal human uroepithelium were established, exposed to a range of 60Co gamma-ray doses from 0.1-5 Gy and grown for 14 days. Expression of Myc, p53 and Bc12 proteins in the epithelial cells which grew from irradiated explants was measured in situ using immunocytochemistry. The results show that overexpression of Bc12 with low Myc expression correlated with resistance to radiation as shown by the extent of growth detected on day 14. Strong staining for Myc coupled with low or absent Bc12 expression generally correlated with radiosensitivity, although the level of p53 of the culture was critical in these cases. None of the proteins on their own correlated with radiation response. What appeared to be critical was the balance of cells expressing Bc12 and Myc proteins. Building on the results presented in a previous paper which showed a division of cultures from patients into those showing monotonic and nonmonotonic responses, this study presents results for explant cultures from a greater number of patients and attempts to characterize the profile of expression of the above proteins in the uroepithelium of these patients. It shows that high Bc12/Myc ratios were found in cultures which showed a non-monotonic and resistant dose response. Where Myc was the dominant protein in the culture postirradiation, a radiosensitive and monotonic response tended to occur. Since the proteins are being detected in the distant progeny of irradiated cells, it is likely that changes induced by radiation in the cell population are stable. The measurement of these two proteins can be made in cultured biopsy material and may therefore have predictive value in radiotherapy and radiation protection. Both normal and tumor biopsies from bladder mucosa showed similar correlations between Bc12/Myc ratios and growth postirradiation.

Primary Explants of Human Uroepithelium Show an Unusual Response to Low-Dose Irradiation with Cobalt-60 Gamma Rays

Recent results using very low doses of radiation have suggested that there is a hypersensitive region where cultures show an enhanced level of cell killing leading to a non-monotonic survival curve. This effect has been observed at doses below 2 Gy in mammalian systems and at much higher doses in insect cells. In this paper we report observation of the effect in primary human uroepithelial cell cultures. The effect was measured using a postirradiation proliferation assay where irradiated explants of standard size were allowed to proliferate for 14 days after exposure to 60Co gamma irradiation. By 14 days the majority of cultures derived from explants irradiated with 2-5 Gy showed little evidence of growth inhibition and cell numbers approached or even exceeded those obtained in the controls. There was, however, a significant reduction in cell number and growth rate in all cultures exposed to doses lower than 1 Gy. Oncoprotein (p53, c-myc, bcl-2, p21 ras) and EGFR expression were also measured in these cultures and were significantly increased. Morphological evidence of apoptosis was present in all irradiated cultures at 4 h after exposure, but this persisted for longer periods in cultures exposed to low doses.

Proliferation of Normal and Malignant Human Epithelial Cells Post Irradiation

Fragments of human oesophageal mucosa, urothelium, squamous and adenocarcinoma of the oesophagus and carcinoma of the bladder have been plated in culture and irradiated. The cells growing from the explanted tissues have then been studied for four weeks post irradiation to assess the overall rate of growth from the irradiated explants and the fraction of proliferating cells. The results show that when using cell number as an endpoint it is possible to derive growth curves from this type of data which permit a doubling time to be obtained for the cell population surviving different doses. In an attempt to determine the proliferating fraction of the cell population, cultures were labelled at appropriate intervals with tritiated thymidine and were also stained with Ki-67 antiproliferating antigen. The results show an interesting relationship between the dose response obtained for cell labelling with tritiated thymidine and area of cellular outgrowth. Ki-67 staining when used carefully and analysed as described was a useful indicator of proliferating cells. The results provide a means of determining the post irradiation growth potential of fragments of tissue from human organs and may be important for determined overall response of the tumour bulk to proposed treatment.

Enhanced prostaglandin synthesis after ultraviolet injury is mediated by endogenous histamine stimulation. A mechanism for irradiation erythema.

Acute ultraviolet light B (UVB) injury is associated with dermal mast cell histamine release. The possibility that histamine-stimulated prostaglandin (PG) synthesis could be a mechanism for irradiation erythema was therefore examined using human skin explants. Explants responded to UV irradiation (120 mJ/cm2) with a fivefold increase in synthesis of prostaglandins E2, F2 alpha and 6-keto PGF1 alpha. Incubating explants with the H1 antihistamines brompheniramine (50 microM) or pyrilamine (30 microM) inhibited PG release from irradiated explants 63 +/- 4.9% (mean +/- SEM) 6 h after UV exposure. Antihistamines did not affect PG synthesis in control explants. Irradiation increased the histamine concentration in explant conditioned medium only 50% over basal values, suggesting that irradiation enhanced histamine responsiveness. Explants were therefore incubated with exogenous histamine. In irradiated explants, PG synthesis was stimulated threefold by 3 microM histamine. Unirradiated explants' PG synthesis was unaffected by histamine. Enhanced histamine sensitivity was also examined in epidermal cell cultures. In irradiated cultures, histamine sensitivity was again markedly potentiated: as little as 1 microM histamine stimulated significant PGE2 release and the response to 10-30 microM histamine was increased six to eight times compared with that of unirradiated cultures. These studies demonstrate that endogenous histamine stimulates PG synthesis in human skin after UV injury by potentiation of histamine-induced prostaglandin release. Potentiated agonist responses induced by UV exposure may contribute to the effects of UVB irradiation injury and in particular to irradiation erythema.

Somatic Embryogenesis in Maize and Comparison of Genetic Variability Induced by Gamma Radiation and Tissue Culture Techniques

AbstractSixteen inbred lines and one hybrid of manse were tested for their capability of somatic embryogenesis, and fully developed plants could be regenerated, from ten inbred, lines. The highest frequency of plant regeneration was expressed in the inbred line CHI 31, and when this line was crossed with a recalcitrant, non‐regenerating line, the F1 and BC hybrids were regenerable. The results of reciprocal crosses demonstrated that dominant nuclear genes and cytoplasmic factors are primarily responsible for the heritable determination of embryogenic callus proliferation and in vitro regeneration of maize plants.Somaclonal and radiation‐induced variability was studied in maize to assess their nature and potential contribution to plant breeding., The inbred line CHI 31 possessing a high in vitro capacity of somatic embryo formation was used as experiments.] material. CHI 31 plants were selfed and twelve‐day old zygotic embryos irradiated with 60Co gamma radiation in situ. Mature caryopses were harvested and assigned as M1 material. In another series, immature zygotic embryos (size 1.2—1.5 mm) were cultured in vitro on N‐6 medium supplemented with 2,4‐D (2.5 μM), and somatic embryos regenerated into plants; these were transplanted into soil and self‐pollinated. Regenerants from non‐irradiated cultures were grown as R1 generation, while regenerants from irradiated explants were considered as M1R1 generation.The genetic variability was evaluated in the M2, R2 and M2R2 generations, respectively, and compared with a non‐treated seed control. Irradiation induced a variety of chlorophyll and morphological variants segregating in the M; generation; however, the frequency of deviant types obtained in the R: generation (somaclonal variation) was significantly exceeding the one derived from the M2 populations. The combination of expert irradiation and in vitro regeneration was most effective for the manifestation of chlorophyll and morphological o if types in the M2R2 generation, and increased drastically the frequency of early flowering variants. Differences in the segregation patterns of mutant phenotypes amonsister somaclones in the R3 and M3R3 generations indicate a different genetic basis, of plants originating from the same explant. This phenomenon suggests a mutational sectoring of the callus during culture. Radiation induced and somaclonal variation exerted a similar spectrum of chlorophyll and morphological deviants.

Effets de rayons γ 60Co sur des vitroplants haploïdes et diploïdes de Gerbera jamesonii

Gerbera jamesonii ‘Super Gerbera’ explants were irradiated in vitro with 60Co γ-rays (8–100 Gy). Some received a single dose, while others received two or three at successive multiplication cycles. Several parameters were measured after 6 weeks of in vitro culture, to compare the effects of irradiation on haploid and diploid clones. Productivity is linked to the ploidy level, whereas total growth is not. Total growth inhibition is proportionate to the given dose. Callus and plantlet growth are either inhibited or stimulated, depending on the type of treatment. Callus growth is stimulated by low doses, which induce the transformation of apices into calli; treatment of successive cycles with low doses (20 and 30 Gy for diploids, 10 and 15 Gy for haploids) ensures better plantlet development. High doses inhibited growth of calli and plantlets up to the 50% level. Variations in morphology and in chlorophyll and anthocyanin content were observed among haploid and diploid irradiated explants. Plantlet production is a good criterion for an overall evaluation. Optimal dosage to ensure 50% growth and multiplication rates is estimated at 26 Gy for diploid clones and 16 Gy for haploid clones.

Phytochrome Regulation of the Response to Exogenous Gibberellins by Epicotyls of Vigna sinensis

The elongation rate of cowpea epicotyls from whole cowpea (Vigna sinensis) seedlings and derooted and debladed plants (explants) increased after the main light period (8-hour duration) was extended with either continuous low intensity tungsten light or brief (5 minutes) far-red (FR) irradiation. This end-of-day FR effect was reversed by red (R) irradiation suggesting the involvement of phytochrome. These results confirm and extend those obtained previously with other species. Localization studies indicate the epicotyl to be the site of the photoreceptor. Treatment of cowpea seedlings with paclobutrazol, a gibberellin (GA) biosynthetic inhibitor, abolished the FR promoted epicotyl elongation, indicating a role for GAs in this process. There was no significant difference in epicotyl elongation rates of R plus FR irradiated explants treated with GA(1) or GA(20) and R irradiated explants treated with GA(1). However, R irradiation inhibited subsequent epicotyl elongation of GA(20) treated explants. Moreover, the observation, using GC-MS, that GA(1) and GA(20) are native GAs in cowpea lends support to the concept that phytochrome may control the conversion of endogenous GA(20) to GA(1) in cowpea.

Radiation-induced low-temperature tolerant solid mutants of Chrysanthemum morifolium Ram.

From irradiated explants of the chrysanthemum cultivar Parliament solid non-chimeric low-temperature tolerant mutants were obtained. Some of the mutants flowered approx. 2 weeks earlier than control plants at a temperature of 12°C D/N and looked otherwise identical to the original genotype. The significance of it is discussed, a.o. in relation to attempts in developing selection methods at cell level for such characteristics.

The action of low-intensity pulsed ultrasound on amphibian embryonic tissues.

Amphibian embryos at different stages of development, and ectomesodermal explants (small pieces of embryonic tissue extirpated from lateral parts of the embryos just after neurulation), have been irradiated with continuous and pulsed ultrasound at a frequency of 0.88 MHz, spatial and temporal average intensities being within the range 0.025-0.1 W cm2. Ultrasound pulse repetition frequencies were varied within 10-1000 Hz, duty factor being constant and equal to 0.5. Destruction of embryonic tissue as an immediate effect, and interruption or distortion of development as delayed (24 hr) effects, were registered. Significant damaging effects of non-cavitational and non-thermal origin and dependent on pulse repetition frequencies, were observed. Highly frequency-dependent resonance-type effects were obtained for irradiated explants. The maximum effect on Rana temporaria explants was obtained at pulse repetition frequencies of 10-20 Hz and on Xenopus laevis explants at 110-130 Hz.

The Effects of γ Irradiation on Embryonic Bone and Cartilage in Vitro

In this study tissue culture methods were used to investigate the effects of ionizing irradiation on bone and cartilage of calvaria and radii of 18-day-old embryonic rats (resp. mice). The radii were studied histologically after 2 days of culture following the irradiation. A number of changes were observed. Decrease in growth of the bone collar and of the total rudiment together with an inhibition of the development of the marrow cavity could be established after a radiation absorbed dose of 500-1000 rad. The differences in length and sometimes in thickness of the bone collar between the control and the irradiated explants showed an inhibitory effect of irradiation on bone formation. This finding was corroborated by biochemical experiments with embryonic calvaria in which we found an inhibition of ${}^{14}{\rm C}\text{-proline}$ incorporation into the insoluble collagen fraction. In contrast with these findings we often found a stimulation of ectopic matrix deposition in the primiti...

Abscission of Phaseolus and Impatiens Explants

Stem-petiole explants from the lower pulvinus of the primary leaves of Phaseolus vulgaris L. cv. Red Kidney and from Impatiens sultani Hook cv. Scarlet Baby were exposed to varying dosages of gamma-radiation. With bean, irradiation of 175 to 525 kiloroentgens (kR) significantly accelerated the onset of abscission with a maximum response at 175 to 280 kR. Higher dosages (beginning at 600-700 kR) usually prevented abscission. With Impatiens, 18 to 35 kR significantly accelerated both the onset of abscission and possibly the initial abscission rate; 350 kR cut the time to 100% abscission in half and substantially accelerated the initial abscission rate. Inhibition of abscission in Impatiens was not possible with the available dose rate (35 kR/hour).The accelerating effect of ionizing radiation was apparently due to primarily a reduction in auxin activity, thus accelerating the shift from the relatively ethylene-insensitive stage I to the ethylene-sensitive stage II. In vitro irradiation of exogenous auxin in agar substantially reduced the abscission-delaying effects of the auxin. The low dosage accelerating effects of the ionizing radiation were negated by exogenous auxin application, either prior to or following irradiation. With prior auxin application, a higher concentration of auxin was required to achieve the same delaying effects as with application following irradiation. Apparently due to this reduction in auxin activity, irradiation hastened sensitivity to exogenous ethylene.The accelerating effects of ionizing radiation may be secondarily due to increased levels of endogenous ethylene. With bean tissues, exposures of 262 through 1500 kR of gamma radiation increased the measurable levels of endogenous ethylene released into the ambient air within 2 or 3 days after irradiation.The results of actinomycin D treatment suggested that required de novo RNA synthesis occurred about 4 hours earlier in those Impatiens explants that had received 105 kR gamma radiation than in the nonirradiated controls.An increased mobilization of Ca from the abscission zone may be a contributing factor in the radiation-induced acceleration of abscission. For both bean and Impatiens, exogenous Ca inhibited abscission of irradiated and nonirradiated explants. In addition, a more concentrated Ca solution was required to inhibit abscission of irradiated bean explants, as compared to the nonirradiated controls.

Müllerian-inhibiting activity of human fetal testicular tissue deprived of germ cells by in vitro irradiation.

Extract: Fragments of testicular tissue from seven human fetuses were explanted in three organ-culture dishes for each fetus. One remained unirradiated while the others were submitted to, respectively, 500 and 700 rads of 6 0Co γ-rays. After an interval of 1–2 weeks, the mullerian-inhibiting activity of the explants was tested using the 14 0.5-day-old rat miillerian duct as end organ and the number of germ cells present in 50 orthogonal sections of seminiferous tubules was determined on a minimum of 10 histologic sections. In control explants, only 12.9% of the original number of germ cells were normal after 1–2 weeks in vitro. The mortality of germ cells was increased by γ-rays; only 7% of their original number surviving 500 rads, and 3% 700 rads. Mullerian-inhibiting activity of both control and irradiated explants was normal. Speculation: Mullerian-inhibiting activity of human fetal testicular explants is not affected either by in vitro survival or by γ-irradiation, although both these procedures significantly deplete the germ cell population of the seminiferous tubules, only 3% surviving exposure to 700 rads. If germ cells were involved in the secretion of the antimullerian hormone, such a drastic reduction of their number would certainly be reflected by an impairment of the mullerian-inhibiting activity of the irradiated explants. As this is not the case, and as it has been demonstrated previously that the antimullerian hormone is synthesized by seminiferous tubules and not by interstitial tissue, we speculate that Sertoli cells are the source of the hormone.