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Pituitary-Gonadal Function Before and After Vasectomy
Vol. 23. No. 10. October 1972 Printed in U.S.A.
FERTILITY AND STERILITY
Copyright © 1972 by The Williams & Wilkins Co.
PITUITARY-GONADAL FUNCTION BEFORE AND AFTER VASECTOMY RALPH G. WIELAND, M.D., MARVIN C. HALLBERG, B.S., ELINOR M. ZORN, M.S., DAVID E. KLEIN, M.D., AND SANFORD S. LURIA, M.D. Department of Medicine, Saint Luke's Hospital and Case Western Reserve University School of Medicine, and the Division of Urology, Saint Luke's Hospital, Cleveland, Ohio 44104
Bilateral vasectomy is becoming an increasingly popular method for controlling family size. Questions have been raised concerning possible harmful emotional and organic consequences. There is exceedingly little substantive data with which to answer these serious questions. The present report contains information on pituitary-gonadal function prior to vasectomy and at the time of azoospermia during the followup period. MATERIALS AND METHODS
Eight males undergoing elective vasectomy are the subjects of this report. The age range was 29-49 years (mean 40.1). They had from 2-5 children. The patients were studied on the day of vasectomy and the studies were repeated from 28-92 days postoperatively (mean 56 days). At the time of the postoperative study they all demonstrated azoospermia on seminal fluid analysis. Preoperative blood samples were obtained in the morning, whereas the postoperative samples were drawn randomly during the day. The serum was separated and frozen until the time of analysis. The serum concentrations of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were measured by doubleantibody immunoassays as previously described utilizing 1125.1 Batch 3 antiserum was utilized for the FSH assay. The gonadotropin samples on each individual were measured in the same assay to avoid interassay variation. They are Received March 24, 1972; revised May 10, 1972.
expressed in milli-international units per milliliters in terms of the 2ndIRP-HMG. Serum from a hypophysectomized individual assayed six times has a mean concentration for FSH of 2.8 mI.U./ml. and LH of 2.6 mI.U./ml. The immunoassay for testosterone previously described 2 has undergone certain modifications to further improve interassay variation and to improve sensitivity. Thus it will be described in detail. For normal males serum samples of 0.5 ml. are taken in duplicate. They are diluted to 1.0 ml. with distilled water. The samples are extracted with 5.0 ml. of fresh diethyl ether. A 4.0-ml. aliquot of this extract is transferred to a 10 x 75 mm. glass culture tube and dried under a stream of nitrogen in a 35° C. water bath. The walls are rinsed down and the ether rinse evaporated. In an attempt to equalize the amount of ether extractable material other than testosterone in the standards, an extract of pooled female serum is combined with the standards. The extraction is performed in the ratio 5 ml. of ether to 2 ml. of serum. One milliliter of the ether extract is pipetted into each tube containing standard (each point is run in duplicate). Four milliliters of the extract (equivalent to 1.6 ml. of the serum pool) are run in duplicate for assay of the pool. The additions of 1% human serum albumin and tracer testosterone-I, 2-H 3 are performed as previously described. Recently we have been treating the testosterone-3-bovine serum albumin antibody with a 0.4% (6,9-diamino-2-ethoxy-
779
780
Vol. 23
PITUITARY-GONADAL FUNCTION
TABLE 1. Levels of Testosterone and Gonadotropins in Males before and after Vasectomy After
Before
Patients
LH
FSH
T*
LH
FSH
ng./IOO mi.
mI.U./ml.
mI.U./ml.
ng./IOO mi.
m1.U./ml.
mI.U./ml.
721 458 428 520 490 381 472 652
17.2 9.5 11.4 16.0 6.2 6.7 6.0 7.3
7.4 6.6 14.6 6.1 16.4 6.2 4.2 12.0
634 366 389 366 415 315 268 828
19.7 11.6 25.2 15.2 5.7 8.7 7.2 11.5
9.6 7.9 11.0 13.7 11.1 24.3 9.9 11.7
515 ± 115 667 ± 248 n = 17
10.0 ± 4.5 10.0 ± 3.2 n = 17
9.2 ± 4.5 8.7 ± 2.9 n = 14
T*
L.R W.L. F. B. T. O. R G. A.W. R P. RR Means ± S.D. Normal males
448
±
188
13.1
±
6.6
12.4
±
5.1
* T, testosterone.
acridine lactate) aqueous solution in a ratio of 1 ml. of serum to 4 ml. of the solution. They are mixed, allowed to stand for 15 min., and then centrifuged. The supernatant is transferred to another centrifuge tube and 100 mg. of Norit A charcoal added and mixed. This is centrifuged after standing for 15 min. The addition of charcoal is repeated at least twice more until the supernatant is clear. It is now in a 1: 5 aqueous dilution. It is then further diluted to a final concentration of 1: 500 in phosphate-buffered saline at the time of assay. The addition of the goat anti-rabbit -y-globulin and the liquid-scintillation counting are unchanged. 3 The standard curve is plotted on semilogarithmic paper. The concentration of testosterone in the 1.6 ml. of female pool is read from the standard curve and then appropriately calculated to give the amount of testosterone in the 0.4 ml. used with the standards. The standard curve is then replotted correcting for the added serum. The testosterone concentration of the unknown samples is then read from the corrected curve and expressed as nanograms/lOO ml. The interassay variation was determined by repeatedly assaying a male and a female
pool. The male pool assayed 10 times gave a mean value of 584 ng./l00 ml. ± 14 (S.E.) and the female pool assayed 13 times gave a mean value of 79 ng./l00 ml. ± 3 (S.E.). The precision of the assay was checked further by adding 0.5, 1.0, and 3.0 ng. of testosterone to a previously assayed pool of mixed male and female sera. Three assays run in duplicate gave the following respective recovery values: 0.52, 1.04, and 2.73 ng.; 0.50, 0.99, and 3.13 ng.; 0.49, 0.98, and 2.75 ng. RESULTS AND SUMMARY
The results of the control and postvasectomy values for FSH, LH, and testosterone are shown in Table 1. The mean FSH values are lower than those previously reported from this laboratory. I, 3 This is because of the use of Batch 3 antiserum in the current study. No significant hormonal changes occurred during the period of observation. This report represents the first study concerned with pituitary-gonadal relationships following vasectomy. The control testosterone levels are somewhat lower than those we have observed in a random
October 1972
781
WIELAND ET AL.
normal adult male population, but they are not significantly different. Our mean adult levels are similar to those obtained by other investigators utilizing similar or different technics. 4 , 5 We can conclude from our data that no significant shortterm changes in pituitary gonadotropins or testosterone occur following bilateral vasectomy. Changes observed in certain individuals are probably based on spontaneous variations which we have observed in the peripheral levels of gonadotrophins and testosterone with frequent (15 min.) sampling. 6 Acknowledgments. Certain materials used in the gonadotropin assay were provided by the National Pituitary Agency, Endocrine Study Section, and the National Institute of Arthritis and Metabolic Diseases.
REFERENCES 1. GUEVARA, A., VIDT, D., HALLBERG, M. C., ZORN, E. M., POHLMAN, C., AND WIELAND, R. G. Serum gonadotropin and testosterone levels in uremic males undergoing intermittent dialysis Metabolism 18:1062, 1969. 2. CHEN, J. C., ZORN, E. M., HALLBERG, M. C., AND WIELAND, R. G. Antibodies to testosterone-3-bovine serum albumin, applied to assay of serum 1713-01 androgens. Clin Chem 17:581, 1971. 3. WIELAND, R. G., CHEN, J. C., ZORN, E. M., AND HALLBERG, M. C. Correlation of growth, pubertal staging, growth hormone, gonadotropins and testosterone levels during the pubertal growth spurt in males. J Pediat 79:999, 1971. 4. FURUYAMA, S., MAYES, D. M., AND NUGENT, C. A. A radioimmunoassay for plasma testosterone. Steroids 16:415, 1970. 5. RIVAROLA, M. A., AND MIGEON, C. J. Determination of testosterone and androst-4-ene,3, 17-diene concentration in human plasma. Steroids 7:103, 1966. 6. WIELAND, R. G., AND HALLBERG, M. C.Unpublished observations.
FERTILITY AND STERILITY
Copyright © 1972 by The Williams & Wilkins Co.
PITUITARY-GONADAL FUNCTION BEFORE AND AFTER VASECTOMY RALPH G. WIELAND, M.D., MARVIN C. HALLBERG, B.S., ELINOR M. ZORN, M.S., DAVID E. KLEIN, M.D., AND SANFORD S. LURIA, M.D. Department of Medicine, Saint Luke's Hospital and Case Western Reserve University School of Medicine, and the Division of Urology, Saint Luke's Hospital, Cleveland, Ohio 44104
Bilateral vasectomy is becoming an increasingly popular method for controlling family size. Questions have been raised concerning possible harmful emotional and organic consequences. There is exceedingly little substantive data with which to answer these serious questions. The present report contains information on pituitary-gonadal function prior to vasectomy and at the time of azoospermia during the followup period. MATERIALS AND METHODS
Eight males undergoing elective vasectomy are the subjects of this report. The age range was 29-49 years (mean 40.1). They had from 2-5 children. The patients were studied on the day of vasectomy and the studies were repeated from 28-92 days postoperatively (mean 56 days). At the time of the postoperative study they all demonstrated azoospermia on seminal fluid analysis. Preoperative blood samples were obtained in the morning, whereas the postoperative samples were drawn randomly during the day. The serum was separated and frozen until the time of analysis. The serum concentrations of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were measured by doubleantibody immunoassays as previously described utilizing 1125.1 Batch 3 antiserum was utilized for the FSH assay. The gonadotropin samples on each individual were measured in the same assay to avoid interassay variation. They are Received March 24, 1972; revised May 10, 1972.
expressed in milli-international units per milliliters in terms of the 2ndIRP-HMG. Serum from a hypophysectomized individual assayed six times has a mean concentration for FSH of 2.8 mI.U./ml. and LH of 2.6 mI.U./ml. The immunoassay for testosterone previously described 2 has undergone certain modifications to further improve interassay variation and to improve sensitivity. Thus it will be described in detail. For normal males serum samples of 0.5 ml. are taken in duplicate. They are diluted to 1.0 ml. with distilled water. The samples are extracted with 5.0 ml. of fresh diethyl ether. A 4.0-ml. aliquot of this extract is transferred to a 10 x 75 mm. glass culture tube and dried under a stream of nitrogen in a 35° C. water bath. The walls are rinsed down and the ether rinse evaporated. In an attempt to equalize the amount of ether extractable material other than testosterone in the standards, an extract of pooled female serum is combined with the standards. The extraction is performed in the ratio 5 ml. of ether to 2 ml. of serum. One milliliter of the ether extract is pipetted into each tube containing standard (each point is run in duplicate). Four milliliters of the extract (equivalent to 1.6 ml. of the serum pool) are run in duplicate for assay of the pool. The additions of 1% human serum albumin and tracer testosterone-I, 2-H 3 are performed as previously described. Recently we have been treating the testosterone-3-bovine serum albumin antibody with a 0.4% (6,9-diamino-2-ethoxy-
779
780
Vol. 23
PITUITARY-GONADAL FUNCTION
TABLE 1. Levels of Testosterone and Gonadotropins in Males before and after Vasectomy After
Before
Patients
LH
FSH
T*
LH
FSH
ng./IOO mi.
mI.U./ml.
mI.U./ml.
ng./IOO mi.
m1.U./ml.
mI.U./ml.
721 458 428 520 490 381 472 652
17.2 9.5 11.4 16.0 6.2 6.7 6.0 7.3
7.4 6.6 14.6 6.1 16.4 6.2 4.2 12.0
634 366 389 366 415 315 268 828
19.7 11.6 25.2 15.2 5.7 8.7 7.2 11.5
9.6 7.9 11.0 13.7 11.1 24.3 9.9 11.7
515 ± 115 667 ± 248 n = 17
10.0 ± 4.5 10.0 ± 3.2 n = 17
9.2 ± 4.5 8.7 ± 2.9 n = 14
T*
L.R W.L. F. B. T. O. R G. A.W. R P. RR Means ± S.D. Normal males
448
±
188
13.1
±
6.6
12.4
±
5.1
* T, testosterone.
acridine lactate) aqueous solution in a ratio of 1 ml. of serum to 4 ml. of the solution. They are mixed, allowed to stand for 15 min., and then centrifuged. The supernatant is transferred to another centrifuge tube and 100 mg. of Norit A charcoal added and mixed. This is centrifuged after standing for 15 min. The addition of charcoal is repeated at least twice more until the supernatant is clear. It is now in a 1: 5 aqueous dilution. It is then further diluted to a final concentration of 1: 500 in phosphate-buffered saline at the time of assay. The addition of the goat anti-rabbit -y-globulin and the liquid-scintillation counting are unchanged. 3 The standard curve is plotted on semilogarithmic paper. The concentration of testosterone in the 1.6 ml. of female pool is read from the standard curve and then appropriately calculated to give the amount of testosterone in the 0.4 ml. used with the standards. The standard curve is then replotted correcting for the added serum. The testosterone concentration of the unknown samples is then read from the corrected curve and expressed as nanograms/lOO ml. The interassay variation was determined by repeatedly assaying a male and a female
pool. The male pool assayed 10 times gave a mean value of 584 ng./l00 ml. ± 14 (S.E.) and the female pool assayed 13 times gave a mean value of 79 ng./l00 ml. ± 3 (S.E.). The precision of the assay was checked further by adding 0.5, 1.0, and 3.0 ng. of testosterone to a previously assayed pool of mixed male and female sera. Three assays run in duplicate gave the following respective recovery values: 0.52, 1.04, and 2.73 ng.; 0.50, 0.99, and 3.13 ng.; 0.49, 0.98, and 2.75 ng. RESULTS AND SUMMARY
The results of the control and postvasectomy values for FSH, LH, and testosterone are shown in Table 1. The mean FSH values are lower than those previously reported from this laboratory. I, 3 This is because of the use of Batch 3 antiserum in the current study. No significant hormonal changes occurred during the period of observation. This report represents the first study concerned with pituitary-gonadal relationships following vasectomy. The control testosterone levels are somewhat lower than those we have observed in a random
October 1972
781
WIELAND ET AL.
normal adult male population, but they are not significantly different. Our mean adult levels are similar to those obtained by other investigators utilizing similar or different technics. 4 , 5 We can conclude from our data that no significant shortterm changes in pituitary gonadotropins or testosterone occur following bilateral vasectomy. Changes observed in certain individuals are probably based on spontaneous variations which we have observed in the peripheral levels of gonadotrophins and testosterone with frequent (15 min.) sampling. 6 Acknowledgments. Certain materials used in the gonadotropin assay were provided by the National Pituitary Agency, Endocrine Study Section, and the National Institute of Arthritis and Metabolic Diseases.
REFERENCES 1. GUEVARA, A., VIDT, D., HALLBERG, M. C., ZORN, E. M., POHLMAN, C., AND WIELAND, R. G. Serum gonadotropin and testosterone levels in uremic males undergoing intermittent dialysis Metabolism 18:1062, 1969. 2. CHEN, J. C., ZORN, E. M., HALLBERG, M. C., AND WIELAND, R. G. Antibodies to testosterone-3-bovine serum albumin, applied to assay of serum 1713-01 androgens. Clin Chem 17:581, 1971. 3. WIELAND, R. G., CHEN, J. C., ZORN, E. M., AND HALLBERG, M. C. Correlation of growth, pubertal staging, growth hormone, gonadotropins and testosterone levels during the pubertal growth spurt in males. J Pediat 79:999, 1971. 4. FURUYAMA, S., MAYES, D. M., AND NUGENT, C. A. A radioimmunoassay for plasma testosterone. Steroids 16:415, 1970. 5. RIVAROLA, M. A., AND MIGEON, C. J. Determination of testosterone and androst-4-ene,3, 17-diene concentration in human plasma. Steroids 7:103, 1966. 6. WIELAND, R. G., AND HALLBERG, M. C.Unpublished observations.