Signalling pathways involved in the control of sperm cell volume

Abstract

The ability to maintain cellular volume is an important general physiological function, which is achieved by specific molecular mechanisms. Hypotonically induced swelling results in the opening of K C and Cl K ion channels, through which these ions exit with accompanying water loss. This process is known as regulatory volume decrease (RVD). The molecular mechanisms that control the opening of the ion channels in spermatozoa are as yet poorly understood. The present study investigated pathways of osmo-signalling using boar spermatozoa as a model. Spermatozoa were diluted into isotonic and hypotonic Hepes-buffered saline in the presence or absence of effector drugs, and at predetermined intervals volume measurements were performed electronically. Treatment with protein kinase C (PKC) inhibitors staurosporine, bismaleimide I and bismaleimide X led to dosedependent increases of both isotonic and hypotonic volumes (P!0.05). However, as the isotonic volume was affected more than the hypotonic volume, the kinase inhibitors appeared to improve RVD, whereas activation of PKC with phorbol dibutyrate blocked RVD. The increase in isotonic cell volume induced by bismaleimide X was observed in chloride-containing medium but not in the medium in which chloride was replaced by sulphate, implying that PKC was involved in the control of chloride channel activity, e.g. by closing the channel after volume adjustment. The protein phosphatase PP1/PP2 inhibitors calyculin and okadaic acid increased the isotonic volume only slightly but they greatly increased the relative cell volume and blocked RVD. The activation of RVD processes was found to be cAMP-dependent; incubation with forskolin and papaverine improved volume regulation. Moreover, papaverine was able to overcome the negative effect of protein phosphatase inhibitors. The mechanism of sperm RVD appears to involve (a) alterations in protein phosphorylation/dephosphorylation balance brought about by PKC and PP1 and (b) a cAMP-dependent activating pathway.

Determinants of sperm quality and fertility in domestic species

“…During NVI, the dephosphorylationphosphorylation balance within the controlling signaling system is disturbed, the swelling-stimulated channels which are usually closed under isotonic conditions become activated, ion uptake and coupled water transport takes place, and the cells swell. In this situation, the volume distribution curve is shifted greatly to the right, and the peak is located beyond the reference area appropriate for physiological conditions (Petrunkina et al 2005b(Petrunkina et al , 2007. The changes are accompanied by membrane leakage to propidium iodide.…”

“…Little is known about whether and how phosphorylation processes are linked to the activation of osmosensing regulatory mechanisms in spermatozoa. Work in our laboratory has provided an evidence that Current concepts in sperm function analysis volume regulation under hypertonic conditions is mediated by protein tyrosine kinase-dependent pathways (Petrunkina et al 2005b), while volume regulation following hypotonic stress is activated through protein kinase C-and cAMP-dependent pathways (Petrunkina et al 2007; see Fig. 2B).…”

“…Spermatozoa corresponding to the different populations obviously differ in functional status, most likely in their cytosolic ion content and the functionality of their membrane channels. We have shown recently that the occurrence of additional subpopulations under isotonic conditions is associated with an uncontrolled ion uptake from the external medium and is related to a deficiency in the signaling pathways that control cell volume (Petrunkina et al 2005b(Petrunkina et al , 2007. Because the presence of such peaks is an early sign of progressive instability in the ejaculate, their development is referred to as necrotic volume increase (NVI).…”

PKC Regulation of Gametogenesis and Early Development

“…Although activation of PKC in sperm seems to result in similar developmental fates in a diversity of animals, it is not to be considered a universal activator since PKC activation in fowl sperm blocks the acrosome reaction (Ashizawa et al, 2006). Some level of PKC activity also has been implicated in retaining the proper volume in sperm as it traverses fluids of different osmolarity during its passage from the testis to the oviduct (Petrunkina et al, 2007).…”

Cytometric solutions in veterinary andrology: Developments, advantages, and limitations

“…Using quinine-and 5-nitro-2-(3-phenylpropylamino)-benzoic acid-sensitive channels, respectively, the ions leave in tandem to maintain electroneutrality. The time course of volume regulation is similar in different mammalian sperm species, and the cells are able to regulate their volume almost back to initial levels after primary swelling by protein kinase-dependent pathways (82,83).…”