Pharmacological studies on human sperm motility

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T I P S - F e b r u a r y 1985

Pharmacological studies on human sperm motility There is a g r o w i n g interest in andrological p h a r m a c o l o g y w h i c h investigates the d r u g effects on male reproduction. D r u g - i n d u c e d change of h u m a n s p e r m motility is c o n s i d e r e d to have b o t h theoretical a n d clinical implications. The motility of s p e r m has been o b s e r v e d since the earliest use of microscope b y Leeuwenhoek. In c o n t e m p o r a r y laboratories, such a subjective visual assessment is still the most w i d e l y used m e t h o d for evaluating s p e r m motility. In spite of the fact that a variety of n e w techniques, such as multiple exposure p h o t o g r a p h y 1, laser D o p p l e r spectroscopy 2 a n d turb i d i m e t r y 3 have b e e n d e v e l o p e d in the recent years for i m p r o v i n g the accuracy and precision of motility m e a s u r e m e n t , their value has not yet b e e n established. Caffeine, cAMP, carnitine, arginine, kallikrein and p r o s t a g l a n d i n E2 have all b e e n r e p o r t e d to stimulate h u m a n s p e r m motility2,4; in a d d i t i o n , h u n d r e d s of chemicals have b e e n screened for their s p e r m i m m o b i l i z i n g effect as well as their potential as vaginal contraceptives. Most of these studies, however, should be considered as p r e l i m i n a r y since few p r o v i d e essential pharmacological information such as concentration-resp o n s e curves, m a k i n g c o m p a r i s o n very difficult. In 1981, a t r a n s m e m b r a n e migration m e t h o d was specifically d e s i g n e d as a pharmacological test for c o m p a r i n g the concentrationd e p e n d e n t influence of different drugs on h u m a n s p e r m motility s. It measures the p r o p o r t i o n of s p e r m m o v i n g across the 5 micron pores of a Nuclepore m e m b r a n e from an aliquot of semen to buffer solution. A l t h o u g h it does not record the m o v i n g pattern of ind i v i d u a l sperm, it is a simple, objective, quantitative and reproducible m e t h o d for evaluating the overall change of s p e r m motility in a bulk of ejaculated semen. Because only a small aliquot of s e m e n is r e q u i r e d for each measurement, w i t h i n - s a m p l e comparisons become possible and thus variation d u e to differences a m o n g s e m e n samples can be excluded. Sperm i m m o b i l i z i n g potencies 1985, E l s e v i e r

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of several drugs, including antidepressants 6, antiarrhythmic agents 7, antihistamines s, 13-adrenoceptor blocking agents 9, cannabis 1°, local anesthetics 11, neuroleptics 12, opiates 13 and vaginal contraceptives 14 have b e e n systematically investigated w i t h the transmembrane migration method. These experiments have d e m o n strated that drugs i m m o b i l i z e s p e r m b y acting on sperm m e m brane; lipid solubility, w h i c h d e t e r m i n e s the a d s o r p t i o n of drugs onto cellular m e m b r a n e , correlates w i t h s p e r m i m m o b i l i z i n g potency 9. Sperm i m m o b i l i z a t i o n could b e similar in nature to the m e c h a n i s m b y w h i c h local anesthetics act on nerve fiber 7. Further evidence s u p p o r t i n g the s p e r m m e m b r a n e as the active site for s p e r m i m m o b i l i z i n g agents is that mitochondrial i n h i b i t o r s such as cyanide TM and m i c r o t u b u l a r inhibitors such as colchicine do not i n h i b i t h u m a n s p e r m motility. Thus, h u m a n s p e r m has b e e n suggested as a potential m o d e l for the s t u d y of m e m b r a n e - a c t i v e drugs TM. Screening active i n g r e d i e n t s for vaginal contraceptive activity is a practical application of this research. D-Propranolol, the dextroisomer of propanolol lacks card i o p u l m o n a r y activity b u t possesses potent s p e r m i m m o b i l i z i n g effect and could p o s s i b l y be used for this p u r p o s e 17. A clinical trial involving a total of 127 w o m e n years s h o w e d a favorable result for propranolol as a vaginal contraceptive TM. It should be e m p h a sized that systemic a d m i n i s t r a t i o n of propranolol is unlikely to cause s p e r m i m m o b i l i z a t i o n because the concentration r e q u i r e d is nearly a t h o u s a n d times h i g h e r than that achieved in h u m a n s e m e n after oral dosage 19. A n o t h e r f i n d i n g revealed b y the t r a n s m e m b r a n e migration m e t h o d is that calcium chelators 2° and calcium antagonists 21 stimulate s p e r m motility in ejaculated human semen. In a d d i t i o n , A23187, a calcium i o n o p h o r e that increases intracellular calcium concentration, inhibits sperm motility and antagonizes the stimulatory effect of calcium chelators and calcium antagonists. In6147/85/$02.00

creased intracellular calcium is therefore considered to be detrimental to the motility of ejaculated h u m a n sperm. Caffeine has also been d o c u m e n t e d as a motility stimulator. Its potency in stimulating h u m a n s p e r m is, however, much less than those of calcium chelators and calcium antagonists. For example, 0.05mM EGTA induces a 79.3% increase in s p e r m motility; for a similar a m p l i t u d e of motility stimulation, 7.5 mM of caffeine is required. In spite of the evidence that h u m a n s p e r m motility can either be stimulated or i n h i b i t e d in v i t r o by various pharmacological agents, no orally or parenterally a d m i n i s t e r e d drugs have b e e n identified w h i c h m o d i f y s p e r m motility w i t h o u t concomitantly c h a n g i n g the concentration or the m o r p h o l o g y of sperm. In other words, all currently available drugs that i m p r o v e or i m p a i r male fertility are considered to act on the production, m a t u r a t i o n or passive transport, rather than the active m o v e m e n t of sperm. Sperm motility does not guarantee conception b u t i m m o t i lity precludes it. Pharmacological studies on s p e r m motility have clarified the inter-relationship between calcium ion and s p e r m motility, b u t have not s h o w n w h e t h e r calcium chelators a n d calcium antagonists can b e used for i m p r o v i n g the successful rate of artificial i n s e m i n a t i o n or for the m a n a g e m e n t of subfertile m e n w i t h p o o r s p e r m motility. A n interdisciplinary co-operation is urgently n e e d e d for further exploration in this field. c. Y. HONG Department of Medicine, Veterans General Hospital and National Yang-Ming Medical College, Taipei, Taiwan.

References 1 Makler, A., Makler, E., Itzkovitz, J. and Brandes, J.M. (1980) Fertil. Steril. 33, 624-630 2 Hartmann, R., Steiner, R., Hofmann, N. and Kaufmann, R. (1982) Andrologia 15, 120-134 3 Levin, R. M., Shofer, J and Greenberg, S. H. (1980) Fertil. Steril. 33, 631--635 4 Amelar, R. D., Dubin, L. and Schoenfeld, C. Y. (1980) Fertil. Steril. 34, 197215 5 Hong, C. Y., Chaput de Saintonge, D. M. and Turner, P. (1981) Br. J. Clin. Pharmacol. 11, 385-387 6 Hong, C. Y., Chiang, B. N. and Ku, J. (1984) Arch. Androl. 12, 25-28 7 Hong, C. Y. and Chiang, B. N. (1984) Br.

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T I P S - F e b r u a r y 1985 ]. Clin. Pharmacol. 17, 687~90 8 Thomas, M. and Turner, P. (1983) J. Pharm. Pharmacol. 35, 361-362 9 Hong, C. Y. and Turner, P. (1982) Br. J. Clin. Pharmacol. 14, 269-272 10 Hong, C. Y., Chaput de Saintonge, D.M. and Turner, P. (1982) Human Toxicol. 1,151-154 11 Hong, C. Y. (1982) J. Physiol. (London) 325, 37P 12 Hong, C. Y., Chaput de Saintonge, D. M. and Turner, P. (1982) Eur. J. Clin. Pharmacol. 22, 413-416

13 Zaman, S., Lamb, J. M., Esberger, D. A. and Pearson, R.M. (1984) Br. J. Clin. Pharmacol. 18, 302P 14 Hong, C. Y., Chiang, B. N. and Wu, P. (1983) Clin. Reprod. Fertil. 2, 33-38 15 Hong, C. Y., Chiang, B.N. and Wei, Y.H. (1983) Br. J. Clin. Pharmacol. 16, 487-490 16 Hong, C. Y., Chiang, B. N., Ku, J. and Wei, Y. H. (1984) Human Toxicol. 3, 271277 17 Hong, C. Y., Chaput de Saintonge, D. M. and Turner, P. (1982) Br. J. Clin.

'~tial event of acetylcholine-induced, EDRF-mediated vasodilatation may involve phosphatidylinositol turnover-coupled Ca ~ mobilization'

Endothelium-dependent relaxing factor and calcium In their recent article, Brown and Masters (TIPS, October 1984) 1 briefly discussed Ca 2+ mobilization and p h o s p h o i n o s i t i d e hydrolysis in relation to the relaxation of vascular smooth muscle which occurs as an ' i n h i b i t o r y ' muscarinic response. They m e n t i o n e d that ' p h o s p h o i n o s i t i d e turnover and Ca 2+ m o b i l i z a t i o n . . , may lead to the formation and release of a secondary mediator' by vascular endothelial cells which m i g h t be responsible for the vasodilatation that follows activation of endothelial muscarinic receptors. As this mechanism, which involves a signal termed endothel i u m - d e p e n d e n t relaxing factor(s) (EDRF(s)), has received m u c h attention recently2, some discussion of studies conducted on the possible role of Ca 2+ might be of interest. In this regard, activation of the EDRF m e c h a n i s m by the Ca 2+ionophore A231872,3 could occur by m e a n s of an increase in intracellular Ca 2+ mobilization (i.e., a translocation of Ca 2+ from one b o u n d state to another b o u n d state) in the endothelial cell, or by an increase in Ca 2+ entry into the cell which w o u l d promote intracellular Ca 2+ mobilization. This change in Ca 2+ could activate p h o s p h o l i p a s e A 2 a n d / o r diglyceride lipase, leading to an increased production of free fatty acids (e.g., arachidonate) which could be

substrates for EDRF formation. With A23187 as the stimulus for relaxation, the sequence of events in the endothelial cell might be: mobilization of intracellular Ca 2+ activation of phospholipase A2 a n d / o r diglyceride lipase (two Ca2+-dependent enzymes) --* release of arachidonate ~ synthesis of EDRF(s). The phospholipase C step (phosphoinositide hydrolysis) could be 'bypassed', and phospholipase A2 a n d / o r diglyceride lipase could be directly activated. As higher concentrations of verapamil (10-5 M) or n i f e d i p i n e (5 x 10-7 M) are required to inhibit m a x i m u m methacholine- or A23187-induced endotheliumd e p e n d e n t relaxations of rabbit aortic strips 3 than relaxations due to Ca2+-entry, such effects might be related to interactions of these 'slow channel blockers' with ACh receptors of the endothelial cell and/or with smooth muscle calm o d u l i n 4-6, or with other intracellular Ca2+-dependent processes. Thus, the muscarinic ' i n h i b i tory' (EDRF) m e c h a n i s m appears to be subserved b y some Ca 2+dependent processes, even though the initial event (acetylcholine-receptor interaction) might not be Ca2+-dependent. Perhaps, the most important Ca 2+ - d e p e n d e n t events that follow receptor activation are: release of arachidonate from phospholipids

Pharmacol. 13, 285P 18 Zipper, J., Wheeler, R. G., Potts, D. M and Rivera, M. (1983) Br. Med. J. 287, 1245-1246 19 Mahajan, P., Grech, E.D., Ridgway, E.J., Turner, P. and Pearson, R.M. (1983) Br. J. Clin. Pharmacol. 17, 185P186P 20 Hong, C. Y., Chiang, B. N., Ku, J. and Wei, Y. H. (1984) Lancet i, 460-461 21 Hong, C. Y., Chiang, B. N., Ku, J., Wei, Y. H. and Fong, J. C. Br. J. Clin. Pharmacol. (in press)

of the endothelial cell (by the action of phospholipase A 2 a n d / o r diglyceride lipase) and activation of a protein kinase (e.g., protein kinase C; cGMP-dependent protein kinase) in the smooth muscle cell1,7. These events would eventually favor relaxation of blood vessels. I n h i b i t i o n of the EDRF mechanism by exposure of rabbit aortic strips to Ca2+-free m e d i u m 3 could also be explained by i n h i bition of the a b o v e - m e n t i o n e d processes. In sum, the initial event of acetylcholine-induced, EDRFmediated, vasorelaxation m i g h t involve receptor-linked phosphatidylinositol turnover that is coupled to an increase in intracellular Ca 2+ m o b i l i z a t i o n in the endothelial cell. Ca 2+ seems to be necessary for the synthesis of EDRF(s) in the endothelial cell, and for the activation of a protein kinase in the smooth muscle cell. If EDRF(s) is indeed a molecule(s) that is released from endothelial cells, then, this 'secretory' process, itself, might also be Ca2+-de pendent. F. V. DEFEUDIS

Universit~ Louis Pasteur, Facultd de M~decine, 67084 Strasbourg, Cedex, France.

References 1 B r o w n , J. H. a n d Masters, S. B. (1984) Trends Pharmacol. Sci. 5, 417-419 2 F u r c h g o t t , R. F., C h e r r y , P . D . , Z a w a dzki, J. V. a n d J o t h i a n a d a n , D. (1984) J.

Cardiovasc. Pharmacol. 6, $336-$343 3 Singer, H. A. and Peach, M.J. (1982) Hypertension, 4 (suppl. II), 19-25 4 Fairhurst, A. S., Whittaker, M.L. and Ehlert, F. J. (1980)Biochem. Pharmacol. 29, 155-162 5 Karliner,J. S., Motulsky,H. J., Dunlap,J., Brown, J.H. and Insel, P.A. (1982) J. Cardiovasc. Pharmacol. 4, 515-520 6 Epstein, P. M., Fiss, K., Hachisu, R. and Andrenyak, D.M. (1982) Biochem. Biophys. Res. Commun. 105, 1142-1149 7 Rapoport, R. M. and Murad, F. (1983) J. Cyclic Nucleotide Res. 281-296

1985,ElsevierSciencePublishers B.V.,Amsterdam 0165 6147/85/$O2.00