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Biomedical subjects

A I Frankel

Publications and source records attributed to A I Frankel.

At least 19 recordsLinked to original sources
PubMed1989

The equivocal presence of nuclear androgen binding proteins in mammalian spermatids and spermatozoa.

The presence of nuclear androgen binding proteins measured by nuclear androgen exchange in rat spermatids and spermatozoa was re-examined. Specific binding was observed to be related to less dense contaminating particles when sonicated testes were fractionated by isopcynic centrifugation through a 40-61% Nycodenz linear gradient. No specific binding was observed in a pure preparation of epididymal spermatozoa collected by retrograde perfusion of the cauda epididymidis, even when a nuclear exchange assay of superior sensitivity was used. Contamination could easily be induced by adding prostatic tissue to epididymal spermatozoa prior to sonication. Despite this strong evidence that the measure of nuclear androgen receptors by nuclear exchange in germ cells is artefactual, the persistence of high endogenous concentrations of 5 alpha-dihydrotestosterone and 5 alpha-diols (but not testosterone) in spermatozoa of castrated rats argues for the opposite conclusion.

Androgen-Binding Protein↗
PubMed1989

The testicular response to hemicastration in the male rat cannot be maintained in vitro.

The testicular response to hemicastration (in which testicular vein testosterone from the remaining testis doubles in concentration) was studied in vitro in order to establish whether the response is maintained after testicular tissue is removed from the animal. Decapsulated testes and collagenase-dispersed cells from decapsulated testes of rats were incubated for 24 h after hemicastration and testosterone production was compared with that in tissue collected at the time of surgery. Testosterone concentration in the remaining testis 24 h after hemicastration was significantly (P less than 0.05) higher than in the testis removed at the time of hemicastration, but testosterone production in vitro was similar in both tissues. Apparently the single testis remaining in a hemicastrated rat requires extratesticular support in order to maintain its stimulated state.

Animals↗
PubMed1989

Testes are asymmetric in the testicular hemicastration response of the male rat.

An enquiry was made into variability in the testicular hemicastration response of the mature rat (doubling of testicular vein testosterone concentration from the remaining testis 24 h after hemicastration). The response was shown to be asymmetric, being more reliable and more robust when the left testis was removed first. Apparently the testicular hemicastration response, which has been shown to be neurally controlled, shares the asymmetry which has been reported recently in the neural regulation of unilateral ovariectomy.

Animals↗
PubMed1984

Nuclear androgen binding sites in the male rat. I. Unoccupied sites in the prostate.

A study was made of unoccupied androgen binding sites in the nuclei of ventral prostate glands of male rats. They were measured at 0 degrees C by comparing specific binding of 1 nM [3H]DHT to salt extract of purified nuclei during the first hour with specific binding during both hours. This method was dependent upon demonstrated completion of uptake into unoccupied binding sites within the first hour and linearity of exchange with occupied binding sites during both hours. Unoccupied binding sites were not artefactual. They did not increase if tissue concentration was diluted prior to homogenization, while they decreased if homogenization was delayed after the tissue was minced. They could be occupied, both in vitro (if precharged with at least 1 nM unlabeled DHT) or in vivo, by administering testosterone propionate subcutaneously or by infusing testosterone into the jugular vein. Exposure to a high concentration of unoccupied prostatic cytosolic binding sites (608.4 fmol from castrated rats) as compared to low concentration (29.3 fmol from intact rats) during homogenization had little effect upon nuclear unoccupied binding site concentrations (2.16 fmol/mg DNA vs 2.41 fmol/mg DNA, respectively). In individual rats, concentration of unoccupied nuclear androgen binding sites was 4.61 +/- 1.05 fmol/mg DNA, while total binding site concentration (measured with 10 nM [3H]DHT for 24h at 12 degrees C) was 866 +/- 103 fmol/mg DNA. Unoccupied nuclear binding sites reached their highest concentration in animals 4 months old (15.09 fmol/mg DNA) when animals 21 days through 720 days of age were studied. By use of association and dissociation rates of binding, it was determined that the apparent Kd of nuclear binding sites was 1.11 X 10(-12) M. There were no observed differences between unoccupied and occupied binding sites in steroid specificity or in sedimentation rate in an 8-24% glycerol density gradient. Although no physiological importance can be attributed as yet to unoccupied nuclear androgen binding sites in the prostate, they do provide a convenient comparison with putative androgen binding sites in the nuclei of testicular and epididymal germ cells.

Animals↗
PubMed1984

Nuclear androgen binding sites in the male rat. II. Seminiferous tubules.

A nuclear exchange for androgens in the seminiferous tubules of the mature male rat has been established. The exchange, utilizing technology established on the ventral prostate gland described in the previous paper [7], is based upon LH-20 chromatography of salt extract of purified nuclei. The extract is incubated with [3H]methyltrienolone (R 1881) or [3H]dihydrotestosterone (DHT) at 4 degrees C for 21 h. Bound is separated from free on a hydroxylapatite microcolumn. Nonspecific binding is determined by use of 100-fold molar excess of the unlabeled ligand. Specific binding in an incubation volume of 2 ml containing 1.4 nM [3H]R 1881 was 5.78 +/- 0.57 fmol/mg DNA (mean +/- SE). Specific binding was linearly related to protein concentration. Of the steroids studied, only testosterone competed with DHT in suppression of [3H]DHT (one-sixth the affinity). In 4 experiments by saturation analysis, specific binding was saturable and of high affinity: Kd = 6.1 +/- 0.93 X 10(-10), n = 2.2 +/- 0.41 pM (3.75 +/- 0.77 fmol/mg DNA). A second receptor of low affinity and high capacity was also observed: Kd = 3.28 X 10(-7); n = 161 pM (2.46 pmol/mg DNA). In an analysis of specific binding by ultracentrifugation on an 8-34% glycerol density gradient, 2 radioactively labeled peaks were observed, one at 5.6 S, completely suppressed by 100-fold molar excess of unlabeled steroid, and a second larger peak at 3 S, only partially suppressed. The possibility that marginal binding was due to proteolysis could not be confirmed: (1) Addition of nuclear extract from seminiferous tubules to nuclear extract from the ventral prostate enhanced specific binding in the latter; (2) A 4-h delay in processing of seminiferous tubules did not decrease specific binding; (3) A protease inhibitor (PMSF) did not enhance binding. However, sonication of the testes, retaining only late spermatids and spermatozoa, triple specific binding (16.2 +/- 0.8 fmol/mg DNA). These results confirm our previous reports in which we observed by radioimmunologic analysis the presence of androgen receptors in seminiferous tubules, and in late spermatids and spermatozoa. It is suggested that the sonicated testis is an ideal model in which to study the role of the androgen receptor in the maintenance of spermatogenesis.

Animals↗
PubMed1984

Nuclear androgen binding sites in the male rat. III. Late spermatids and spermatozoa in the testis, with an introduction to epididymal spermatozoa.

Nuclear androgen binding sites were examined in late spermatids (stages 12-19) which resisted sonication of homogenized testes of mature male rats. The measurement of unoccupied binding sites in salt extract of purified spermatid heads by nuclear exchange at -10 degrees C was developed and validated. As in the prostate, unoccupied nuclear androgen binding sites in sonicated testes were in low concentration, were not artefactual, and could be occupied both in vivo and in vitro by exogenous androgens, and uniquely in hemicastrated rats by endogenously compensated androgens in the remaining testis. The properties of occupied binding sites in salt extract of purified spermatid heads (measured by nuclear exchange at 4 degrees C for 48 or more hours with 5 nM [3H]dihydrotestosterone) were almost identical to those of occupied binding sites in nuclei of the ventral prostate, except for their concentration. However, levels of specific binding activity approaching 50 fmol/mg DNA could be expected in salt extract of spermatid pellets, by use of a sulfhydryl reducing agent (dithiothreitol) prior to salt extraction, a protease inhibitor (phenylmethylsulfonyl fluoride) in all buffers, and optimization of the sonication protocol. Nuclear androgen binding sites of sonicated epididymal spermatozoa, collected by retrograde perfusion of the cauda epididymidis, were found to be completely salt-resistant. These binding proteins could be extracted by 0.4 M KCl if dithiothreitol and dihydrotestosterone were incorporated into the sonication buffer, if phenylmethylsulfonyl fluoride was added to all buffers, and if the purified epididymal sperm pellet was treated with sarkosyl, a non-ionic detergent, just before salt extraction. The salt extract of epididymal spermatozoa which were treated as described above contained two binding components: a soluble form which was eluted from hydroxylapatite by increasing concentrations of phosphate buffers, and a non-soluble form, free of DNA, which remained in the hydroxylapatite column, and which contained most of the androgen binding sites. Affinity (Kd) of dihydrotestosterone to the soluble and insoluble fractions of the steroid-binding protein complex was determined to be 0.7 and 0.1 nM, respectively. Salt-resistance of binding proteins in germ cells was shown to develop significantly in the last stages of spermiogenesis.

Animals↗
PubMed1984

Hypophysectomy and hemivasectomy can inhibit the testicular hemicastration response of the mature rat.

Three questions were asked in an attempt to understand how testosterone (T) concentration in the veins of the remaining testis can double within 24 h after hemicastration in the mature rat without a change in plasma luteinizing hormone (LH) levels. These three questions (and their answers) were: 1) Can the testicular hemicastration response occur in hypophysectomized rats? Answer, No. 2) Does LH binding to the testis increase after hemicastration? Answer, No. 3) Is there a neural route to the testis alternate to the superior spermatic plexi? Answer, Yes, apparently there is, since hemivasectomy contralateral to the excised testis partially suppressed the testicular hemicastration response (150.4 +/- 13.2 ng/ml in hemicastrated, sham- hemivasectomized rats [n = 18] vs. 109.4 +/- 11.6 ng/ml in hemicastrated, hemivasectomized rats [n = 18], P less than 0.026). It was concluded that LH was probably necessary to the testicular hemicastration response but that its presence did not provide a mechanism. The response was mediated at least partly through the inferior spermatic nerves associated with the vas deferens. A possible reason, although highly speculative, for failure to previously block the testicular hemicastration response by bilateral denervation of the superior spermatic plexi (Mock and Frankel , 1982) was that during the 12-wk interval between denervation and hemicastration, testicular innervation functionally transferred from the superior spermatic to the inferior spermatic nerves.

Animals↗
PubMed1982

The hemicastrated rat: definition of a model for the study of the regulation of testicular steroidogenesis.

A comprehensive study has been made of the hemicastrated rat from 2 to 12 months of age in order to define what might represent an ideal model in which to study testicular regulation. Although there was no compensatory hypertrophy in the remaining testis of the mature hemicastrated rat, levels of plasma testosterone fell significantly within 4 h after surgery in all age groups older than 3 months, and were restored to normal levels almost immediately, usually within 8 h. There were no significant changes in LH and prolactin, and the significant rise in FSH was sufficiently delayed (2 days or more) to suggest that none of these three hormones was implicated in any obvious way in the compensatory restoration of plasma testosterone levels. Although a single testis was capable of maintaining normal plasma testosterone concentrations, its response to human chorionic gonadotrophin at 24 h after hemicastration was significantly less than that of intact animals, suggesting that the single testis was functioning at near-maximal capacity. The hormonal responses to repetitive blood sampling and to sham-surgery simulated the response to hemicastration remarkably. However, these responses were never statistically significant in within-group analysis, and therefore did not obscure the significant fall of plasma testosterone levels in response to hemicastration. The basic mechanism by which plasma testosterone is restored in the hemicastrated rat is still unknown, but the options have been narrowed.

Animals↗
PubMed1982

A study of the first eight hours in the stabilization of plasma testosterone concentration in the hemicastrated rat.

Plasma levels of testosterone fell within 4 h after hemicastration in the mature male rat, and recovered within 8 h, without a compensatory rise in plasma LH from 5 to 480 min after surgery. Pulsatile release of LH was not observed in any group, suggesting the possibility that its alteration was not stimulating the single testis. Luteinizing hormone releasing hormone (LH-RH) stimulated plasma LH concentration equally in both control and hemicastrated rats, rising more sharply only in sham-hemicastrated rats treated with a low dose of LH-RH. Plasma LH rose significantly at the same time (6h after surgery) in both castrated (in one step) and hemicastrated rats which were castrated (in two steps). Bilateral denervation of the testis did not affect the response of plasma testosterone after hemicastration. There was a remarkably similar response in both plasma LH and testosterone levels to handling, blood collection, anaesthesia, sham-surgery and hemicastration separately or in combination. It was concluded that the response of testosterone to hemicastration was neither related to early changes in plasma LH levels nor to alterations in the hypothalamo-hypophysial axis.

Animals↗
PubMed1982

Response of testosterone to hemicastration in the testicular vein of the mature rat.

A rapid procedure was developed and validated for collection of testicular vein blood from rats. Samples were collected from both testes and the heart within 2-5 min of transfer of the rat to the anaesthesia jar. There was no difference in levels of testosterone in testicular vein serum between successive samples from the same testis or alternate samples from paired testes. Both hemicastration and sham-hemicastration significantly depressed levels of testosterone in testicular vein serum 4 h after surgery. Peripheral serum testosterone levels were significantly depressed after hemicastration but not sham-hemicastration at 4 h after surgery. By 24 h after surgery, testosterone in testicular vein serum was doubled in the hemicastrated rat, and was restored in the sham-hemicastrated rat, thus achieving normal peripheral testosterone levels in both groups. Similar results were observed in bilaterally denervated rats. These rapid changes could not be attributed to serum LH, which maintained steady levels during the course of these experiments. It was concluded that there are two components in the response to hemicastration: a non-specific response within 4 h to sham-surgery or any other disturbance to the animal, which depresses testosterone output sharply, and a specific response within 24 h to hemicastration, which doubles testicular output. It is the non-specific response after sham-surgery which has simulated and obfuscated the response to hemicastration in the male rat.

Animals↗