Skip to main content

Pituitary adenylate cyclase activating polypeptide-mediated intracrine signaling in the testicular germ cells

Abstract

Pituitary adenylate cyclase activating polypeptide (PACAP) is found not only in the brain, but is also abundantly expressed in the testicular germ cells. However, the physiological role of testicular PACAP remains unknown. Autoradiographic studies showed a considerable number of PACAP-specific binding sites in the seminiferous tubules. Immunohistochemistry demonstrated PAC1-receptor (R)-like immunoreactivity (li) in the cytoplasm of round spermatids, aggregated in the acrosome and coexpressed with PACAP-li. Spermatid-enriched fractions were examined for the subcellular localization of PACAP binding sites and PAC1-R-li. The highest levels of PACAP binding sites and PAC1-R-li were found in the cytosolic, followed by the nuclear, and the lowest levels in the membrane fraction. The testicular cytosolic PAC1-R-like protein showed a specific competitive inhibition in the radioreceptor assay for PACAP38 and 27, with a K i of 0.069 nM and 0.179 nM, respectively. The addition of PACAP to the cytosol of spermatids only slightly activated adenylate cyclase, while it markedly stimulated the expression and activation of ERK-type mitogen-activated protein kinase (MAPK). In the PAC1-R-like protein-depleted cytosol, a PAC1-R-specific agonist, maxadilan, did not activate MAPK, but PACAP and VIP still did. Because VPAC2-R, which binds both PACAP and VIP, is expressed in the testis, the findings suggest that cytosolic VPAC2-R-like proteins are also present and coupled to MAPK. The MAPK activation does not seem to require a heterotrimeric G-protein. Because PACAP and its receptors are coexpressed in the cytoplasm of spermatids, endogenous PACAP may directly interact with the cytosolic PAC1-R-like protein without the ligand being released into the extracellular space. This possibility is supported by the observation that cytosolic endogenous PACAP in spermatids was co-immunoprecipitated with the cytosolic PAC1-R. This mechanism may be called “intracrine,” and its physiological significance is discussed.

This is a preview of subscription content, access via your institution.

References

  1. 1

    Miyata, A., Arimura, A., Dahl, R. R., et al. (1989). Biochem. Biophys. Res. Commun. 164, 567–574.

    PubMed  Article  CAS  Google Scholar 

  2. 2

    Miyata, A., Jiang, L., Dahl, R. D., et al. (1990). Biochem. Biophys. Res. Commun. 170, 643–648.

    PubMed  Article  CAS  Google Scholar 

  3. 3

    Arimura, A., Somogyvari-Vigh, A., Miyata, A., Mizuno, K., Coy, D. H., and Kitada, C. (1991). Endocrinology 129, 2787–2789.

    PubMed  CAS  Google Scholar 

  4. 4

    Shioda, S., Legradi, G., Leung, W. C., Nakajo, S., Nakaya, K., and Arimura, A. (1994). Endocrinology 135, 818–825.

    PubMed  Article  CAS  Google Scholar 

  5. 5

    Yanaihara, H., Vigh, S., Kozicz, T., Somogyvari-Vigh, A., and Arimura, A. (1998). Regul. Pept. 78, 83–88.

    PubMed  Article  CAS  Google Scholar 

  6. 6

    Kononen, J., Paavola, M., Penttila, T. L., Parvinen, M., and Pelto-Huikko, M. (1994). Endocrinology 135, 2291–2294.

    PubMed  Article  CAS  Google Scholar 

  7. 7

    Hannibal, J. and Fahrenkrug, J. (1995). Regul. Pept. 55, 111–115.

    PubMed  Article  CAS  Google Scholar 

  8. 8.

    Daniel, P. B. and Habener, J. F. (1997). Abstracts of the Endocrine Society’s 79th Annual Meeting. P3-382, p. 532.

    Google Scholar 

  9. 9

    Gottschall, P. E., Tatsuno, I., Miyata, A., and Arimura, A. (1990). Endocrinology 127, 272–277.

    PubMed  CAS  Google Scholar 

  10. 10

    Arimura, A. (1998). Jpn. J. Physiol. 48, 301–331.

    PubMed  Article  CAS  Google Scholar 

  11. 11

    Shivers, B. D., Gorcs, T. J., Gottschall, P. E., and Arimura, A. (1991). Endocrinology 128, 3055–3065.

    PubMed  CAS  Article  Google Scholar 

  12. 12

    Li, M., Shioda, S., Somogyvari-Vigh, A., Onda, H., and Arimura, A. (1997). Endocrine 7, 183–190.

    PubMed  CAS  Google Scholar 

  13. 13

    Arimura, A. and Shioda, S. (1995). Front. Neuroendocrinol. 16, 53–88.

    PubMed  Article  CAS  Google Scholar 

  14. 14

    Gottschall, P. E., Katsuura, G., Dahl, R. R., Hoffmann, S. T., and Arimura, A. (1988). Biol. Reprod. 39, 1074–1085.

    PubMed  Article  CAS  Google Scholar 

  15. 15

    Krantic, S., Martel, J. C., Weissmann, D., and Quirion, R. (1989). Brain Res. 498, 267–278.

    PubMed  Article  CAS  Google Scholar 

  16. 16

    Zhou, C. J., Shioda, S., Shibanuma, M., et al. (1999). Neuroscience 93, 375–391.

    PubMed  Article  CAS  Google Scholar 

  17. 17

    Litvin, T. N., Kamenetsky, M., Zarifyan, A., Buck, J., and Levin, L. R. (2003). J. Biol. Chem. 278, 15922–15926.

    PubMed  Article  CAS  Google Scholar 

  18. 18

    Arimura, A., Somogyvari-Vigh, A., Weill, C., et al. (1994). Ann. NY Acad. Sci. 739, 228–243.

    PubMed  Article  CAS  Google Scholar 

  19. 19

    Buck, J., Sinclair, M. L., Schapal, L., Cann, M. J., and Levin, L. R. (1999). Proc. Natl. Acad. Sci. USA 96, 79–84.

    PubMed  Article  CAS  Google Scholar 

  20. 20

    Sinclair, M. L., Wang, X. Y., Mattia, M., et al. (2000). Mol. Reprod. Dev. 56, 6–11.

    PubMed  Article  CAS  Google Scholar 

  21. 21

    Luconi, M., Barni, T., Vannelli, G. B., et al. (1998). Biol. Reprod. 58, 1476–1489.

    PubMed  Article  CAS  Google Scholar 

  22. 22

    Lu, Q., Sun, Q. Y., Breitbart, H., and Chen, D. Y. (1999). Arch. Androl. 43, 55–66.

    PubMed  Article  CAS  Google Scholar 

  23. 23

    Wadewitz, A. G., Winer, M. A., and Wolgemuth, D. J. (1993). Oncogene 8, 1055–1062.

    PubMed  CAS  Google Scholar 

  24. 24

    Sun, Q. Y., Breitbart, H., and Schatten, H. (1999). Reprod. Fertil. Dev. 11, 443–450.

    PubMed  Article  Google Scholar 

  25. 25

    Berruti, G. (2000). Exp. Cell Res. 257, 172–179.

    PubMed  Article  CAS  Google Scholar 

  26. 26

    Berruti, G. (2003). Cell. Mol. Biol. (Noisy-le-grand) 49, 381–388.

    CAS  Google Scholar 

  27. 27

    El-Gehani, F., Tena-Sempere, M., and Huhtaniemi, I. (1998). Mol. Cell. Endocrinol. 140, 175–178.

    PubMed  Article  CAS  Google Scholar 

  28. 28

    El-Gehani, F., Tena-Sempere, M., and Huhtaniemi, I. (2000). Biol. Reprod. 63, 1482–1489.

    PubMed  Article  CAS  Google Scholar 

  29. 29

    Krempels, K., Usdin, T. B., Harta, G., and Mezey, E. (1995). Neuropeptides 29, 315–320.

    PubMed  Article  CAS  Google Scholar 

  30. 30

    Daniel, P. B., Kieffer, T. J., Leech, C. A., and Habener, J. F. (2001). J. Biol. Chem. 276, 12938–12944.

    PubMed  Article  CAS  Google Scholar 

  31. 31

    Spengler, D., Waeber, C., Pantaloni, C., et al. (1993). Nature 365, 170–175.

    PubMed  Article  CAS  Google Scholar 

  32. 32

    Monts, B. S., Lee, W. H., Breyer, P. R., et al. (1995). Endocrine 3, 505–510.

    CAS  Google Scholar 

  33. 33

    Berry, S. A., Srivastava, C. H., Rubin, L. R., Phipps, W. R., and Pescovitz, O. H. (1992). J. Clin. Endocrinol. Metab. 75, 281–284.

    PubMed  Article  CAS  Google Scholar 

  34. 34

    Srivastava, C. H., Monts, B. S., Rothrock, J. K., Peredo, M. J., and Pescovitz, O. H. (1995). Endocrinology 136, 1502–1508.

    PubMed  Article  CAS  Google Scholar 

  35. 35

    Mbikay, M., Tadros, H., Ishida, N., et al. (1997). Proc. Natl. Acad. Sci. USA 94, 6842–6846.

    PubMed  Article  CAS  Google Scholar 

  36. 36

    Gray, S. L., Cummings, K. J., Jirik, F. R., and Sherwood, N. M. (2001). Mol. Endocrinol. 15, 1739–1747.

    PubMed  Article  CAS  Google Scholar 

  37. 37

    Hashimoto, H., Shintani, N., Tanaka, K., et al. (2001). Proc. Natl. Acad. Sci. USA 98, 13355–13360.

    PubMed  Article  CAS  Google Scholar 

  38. 38

    Hamelink, C., Tjurmina, O., Damadzic, R., et al. (2002). Proc. Natl. Acad. Sci. USA 99, 461–466.

    PubMed  Article  CAS  Google Scholar 

  39. 39

    Seidah, N. G., Day, R., Hamelin, J., Gaspar, A., Collard, M. W., and Chretien, M. (1992). Mol. Endocrinol. 6, 1559–1570.

    PubMed  Article  CAS  Google Scholar 

  40. 40

    Nakayama, K., Kim, W. S., Torii, S., et al. (1992). J. Biol. Chem. 267, 5897–5900.

    PubMed  CAS  Google Scholar 

  41. 41

    Li, M., Nakayama, K., Shuto, Y., Somogyvari-Vigh, A., and Arimura, A. (1998). Peptides 19, 259–268.

    PubMed  Article  CAS  Google Scholar 

  42. 42

    Basak, A., Toure, B. B., Lazure, C., Mbikay, M., Chretien, M., and Seidah, N. G. (1999). Biochem. J. 343(Pt. 1), 29–37.

    PubMed  Article  CAS  Google Scholar 

  43. 43

    Li, M., Mbikay, M., Nakayama, K., Miyata, A., and Arimura, A. (2000). Ann. NY Acad. Sci. 921, 333–339.

    PubMed  CAS  Article  Google Scholar 

  44. 44

    Li, M., Mbikay, M., and Arimura, A. (2000). Endocrinology 141, 3723–3730.

    PubMed  Article  CAS  Google Scholar 

  45. 45

    Heindel, J. J., Powell, C. J., Paschall, C. S., Arimura, A., and Culler, M. D. (1992). Biol. Reprod. 47, 800–806.

    PubMed  Article  CAS  Google Scholar 

  46. 46

    Rossato, M., Nogara, A., Gottardello, F., Bordon, P., and Foresta, C. (1997). Endocrinology 138, 3228–3235.

    PubMed  Article  CAS  Google Scholar 

  47. 47

    Hueso, C., Carmena, M. J., and Prieto, J. C. (1989). Biochem. Int. 19, 951–958.

    PubMed  CAS  Google Scholar 

  48. 48

    Sreedharan, S. P., Huang, J. X., Cheung, M. C., and Goetzl, E. J. (1995). Proc. Natl. Acad. Sci. USA 92, 2939–2943.

    PubMed  Article  CAS  Google Scholar 

  49. 49

    El-Gehani, F., Tena-Sempere, M., and Huhtaniemi, I. (1998). Endocrinology 139, 1474–1480.

    PubMed  Article  CAS  Google Scholar 

  50. 50

    Re, R. (2003). Am. J. Physiol. Heart Circ. Physiol. 284, H751-H757.

    PubMed  CAS  Google Scholar 

  51. 51

    Cann, M. J., Chung, E., and Levin, L. R. (2000). Dev. Genes Evol. 210, 200–206.

    PubMed  Article  CAS  Google Scholar 

  52. 52

    Lazazzera, B. A. (2001). Peptides 22, 1519–1527.

    PubMed  Article  CAS  Google Scholar 

  53. 53

    McRory, J. and Sherwood, N. M. (1997). Endocrinology 138, 2380–2390.

    PubMed  Article  CAS  Google Scholar 

  54. 54

    Srivastava, C. H., Collard, M. W., Rothrock, J. K., Peredo, M. J., Berry, S. A., and Pescovitz, O. H. (1993). Endocrinology 133, 83–89.

    PubMed  Article  CAS  Google Scholar 

  55. 55

    Fabbri, A., Ciocca, D. R., Ciampani, T., Wang, J., and Dufau, M. L. (1995). Endocrinology 136, 2303–2308.

    PubMed  Article  CAS  Google Scholar 

  56. 56

    Breyer, P. R., Rothrock, J. K., Beaudry, N., and Pescovitz, O. H. (1996). Endocrinology 137, 2159–2162.

    PubMed  Article  CAS  Google Scholar 

  57. 57

    Ohta, M., Funakoshi, S., Kawasaki, T., and Itoh, N. (1992). Biochem. Biophys. Res. Commun. 183, 390–395.

    PubMed  Article  CAS  Google Scholar 

  58. 58

    du Plessis, S. S., Page, C., and Franken, D. R. (2001). Andrologia 33, 337–342.

    PubMed  Article  Google Scholar 

  59. 59

    de Lamirande, E. and Gagnon, C. (2002). Mol. Hum. Reprod. 8, 124–135.

    PubMed  Article  Google Scholar 

  60. 60

    Bellve, A. R. (1993). Methods Enzymol. 225, 84–113.

    PubMed  CAS  Article  Google Scholar 

  61. 61

    Romrell, L. J., Bellve, A. R., and Fawcett, D. W. (1976). Dev. Biol. 49, 119–131.

    PubMed  Article  CAS  Google Scholar 

  62. 62

    Bellve, A. R., Millette, C. F., Bhatnagar, Y. M., and O’Brien, D. A. (1977). J. Histochem. Cytochem. 25, 480–494.

    PubMed  CAS  Google Scholar 

  63. 63

    Bellve, A. R., Cavicchia, J. C., Millette, C. F., O’Brien, D. A., Bhatnagar, Y. M., and Dym, M. (1977). J. Cell Biol. 74, 68–85.

    PubMed  Article  CAS  Google Scholar 

  64. 64

    Gerton, G. L. and Millette, C. F. (1986). Biol. Reprod. 35, 1025–1035.

    PubMed  Article  CAS  Google Scholar 

  65. 65

    Joshi, M. S., Anakwe, O. O., and Gerton, G. L. (1990). J. Androl. 11, 120–130.

    PubMed  CAS  Google Scholar 

  66. 66

    Lessley, B. A. and Garner, D. L. (1983). Biol. Reprod. 28, 447–459.

    PubMed  Article  CAS  Google Scholar 

  67. 67.

    Hinton, R. and Mullock, B. (1997). In: Subcellular fractionation—a practical approach. Graham, J. M. and Rickwood, D. (eds.). Oxford University Press: Oxford, UK, pp. 31–69.

    Google Scholar 

  68. 68.

    Levin, L. R. and Reed, R. R. (1995). J. Biol. Chem. 270, 7573–7579.

    PubMed  Article  CAS  Google Scholar 

Download references

Author information

Affiliations

Authors

Corresponding author

Correspondence to Akira Arimura.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Li, M., Funahashi, H., Mbikay, M. et al. Pituitary adenylate cyclase activating polypeptide-mediated intracrine signaling in the testicular germ cells. Endocr 23, 59–75 (2004). https://doi.org/10.1385/ENDO:23:1:59

Download citation

Key Words

  • Immunohistochemistry
  • testicular germ cells
  • mitogen-activated protein kinase
  • adenylate cyclase
  • soluble adenylate cyclase
  • pituitary adenylate cyclase activating polypeptide
  • vasoactive intestinal peptide
  • cytosolic receptors
  • intracrine
  • prohormone convertase 4 knock-out mouse
  • growth hormone releasing hormone
  • secretin