Simultaneous removal of sperm plasma membrane and acrosome before intracytoplasmic sperm injection improves oocyte activation/embryonic development

Boar spermatozoa were prepared for intracytoplasmic sperm injection (ICSI) by two different treatments to facilitate sperm chromatin decondensation and improve fertilisation rates after ICSI in pigs: spermatozoa were either frozen and thawed without cryoprotectants, or treated with progesterone. Morphological changes of the sperm heads after the treatments were examined and then the activation of oocytes and the transformation of the sperm nucleus following ICSI were assessed. After freezing and thawing, the plasma membrane and acrosomal contents over the apical region of sperm head were lost in all the spermatozoa. Following treatment with 1 mg/ml progesterone, the acrosome reaction was induced in 61% of spermatozoa. After injection of three types of spermatozoa, non-treated spermatozoa and progesterone-treated (i.e. acrosome-reacted) spermatozoa induced oocyte activation, but frozen-thawed spermatozoa induced oocyte activation at a significantly lower rate. Sixty-two per cent of sperm heads remained orcein-negative for 6 h, however, resulting in delayed sperm chromatin decondensation and low male pronuclear formation in the oocytes injected with a non-treated spermatazoon. Since the treatments of freezing and thawing and progesterone for spermatozoa accelerated the initial change in sperm chromatin and the latter treatment induced oocyte activation earlier, it is considered that the delay in oocyte activation and decondensation of sperm chromatin after injection of non-treated spermatozoa is caused by the existence of the sperm plasma membrane. These results show that progesterone treatment efficiently induces the acrosome reaction in boar spermatozoa without destroying their potency for oocyte activation, and the induction of the acrosome reaction results in the promotion of male pronuclear formation after ICSI.

Download Full-text

Intracellular calcium oscillations and activation in horse oocytes injected with stallion sperm extracts or spermatozoa

Reproduction ◽

10.1530/rep.0.1260489 ◽

2003 ◽

pp. 489-499 ◽

Cited By ~ 20

Author(s):

SJ Bedford ◽

M Kurokawa ◽

K Hinrichs ◽

RA Fissore

Keyword(s):

Embryonic Development ◽

Intracellular Calcium ◽

Intracytoplasmic Sperm Injection ◽

Animal Species ◽

Mammalian Species ◽

Calcium Oscillations ◽

Large Animal ◽

Oocyte Activation ◽

Mouse Oocytes ◽

Species Specific

In oocytes from all mammalian species studied to date, fertilization by a spermatozoon induces intracellular calcium ([Ca(2+)](i)) oscillations that are crucial for appropriate oocyte activation and embryonic development. Such patterns are species-specific and have not yet been elucidated in horses; it is also not known whether equine oocytes respond with transient [Ca(2+)](i) oscillations when fertilized or treated with parthenogenetic agents. Therefore, the aims of this study were: (i) to characterize the activity of equine sperm extracts microinjected into mouse oocytes; (ii) to ascertain in horse oocytes the [Ca(2+)](i)-releasing activity and activating capacity of equine sperm extracts corresponding to the activity present in a single stallion spermatozoon; and (iii) to determine whether equine oocytes respond with [Ca(2+)](i) transients and activation when fertilized using the intracytoplasmic sperm injection (ICSI) procedure. The results of this study indicate that equine sperm extracts are able to induce [Ca(2+)](i) oscillations, activation and embryo development in mouse oocytes. Furthermore, in horse oocytes, injection of sperm extracts induced persistent [Ca(2+)](i) oscillations that lasted for >60 min and initiated oocyte activation. Nevertheless, injection of a single stallion spermatozoon did not consistently initiate [Ca(2+)](i) oscillations in horse oocytes. It is concluded that stallion sperm extracts can efficiently induce [Ca(2+)](i) responses and parthenogenesis in horse oocytes, and can be used to elucidate the signalling mechanism of fertilization in horses. Conversely, the inconsistent [Ca(2+)](i) responses obtained with sperm injection in horse oocytes may explain, at least in part, the low developmental success obtained using ICSI in large animal species.

Download Full-text

Sperm plasma membrane damage prior to intracytoplasmic sperm injection: a necessary condition for sperm nucleus decondensation

Human Reproduction ◽

10.1093/oxfordjournals.humrep.a135829 ◽

1995 ◽

Vol 10 (11) ◽

pp. 2960-2964 ◽

Cited By ~ 57

Author(s):

D. Dozortsev ◽

A. Rybouchkin ◽

P. De Sutter ◽

M. Dhont

Keyword(s):

Plasma Membrane ◽

Intracytoplasmic Sperm Injection ◽

Membrane Damage ◽

Sperm Nucleus ◽

Necessary Condition ◽

Plasma Membrane Damage ◽

Sperm Plasma Membrane

Download Full-text

Effect of sperm immobilisation and demembranation on the oocyte activation rate in the mouse

Zygote ◽

10.1017/s0967199499000568 ◽

1999 ◽

Vol 7 (3) ◽

pp. 187-193 ◽

Cited By ~ 33

Author(s):

T. Kasai ◽

K. Hoshi ◽

R. Yanagimachi

Keyword(s):

Plasma Membrane ◽

Plasma Membranes ◽

Human Sperm ◽

Sperm Head ◽

The State ◽

Human Spermatozoa ◽

Oocyte Activation ◽

Activation Rate ◽

Sperm Plasma Membrane ◽

Triton X

To analyse the effect of the state of the sperm plasma membrane on oocyte activation rate following intracytoplasmic sperm injection (ICSI), three types of human and mouse spermatozoa (intact, immobilised and Triton X-100 treated) were individually injected into mouse oocytes. At 30, 60 and 120 min after injection, maternal chromosomes and sperm nuclei within oocytes were examined. Following human sperm injection, the fastest and the most efficient oocyte activation and sperm head decondensation occurred when the spermatozoa were treated with Triton X-100. Intact spermatozoa were the least effective in activating oocytes. Thus, the rate of mouse oocyte activation following human sperm injection is greatly influenced by the state of the sperm plasma membrane during injection. When mouse spermatozoa were injected into mouse oocytes, the rates of oocyte activation and sperm head decondensation within activated oocytes were the same irrespective of the type of sperm treatment prior to injection. We witnessed that live human spermatozoa injected into moue oocytes often kept moving very actively within the ooplasm for more than 60 min, whereas motile mouse spermatozoa usually became immotile within 20 min after injection into the ooplasm. In 0.002% Triton X-100 solution, mouse spermatozoa are immobilised faster than human spermatozoa. These facts seem to suggest that human sperm plasma membranes are physically and biochemically more stable than those of mouse spermatozoa. Perhaps the physical and chemical properties of the sperm plasma membrane vary from species to species. For those species whose spermatozoa have ‘stable’ plasma membranes, prior removal or ‘damage’ of sperm plasma membranes would increase the success rate of ICSI.

Download Full-text

Effects of oocyte activation and treatment of spermatozoa on embryonic development following intracytoplasmic sperm injection in cattle

Theriogenology ◽

10.1016/s0093-691x(97)00369-5 ◽

1997 ◽

Vol 48 (8) ◽

pp. 1265-1273 ◽

Cited By ~ 23

Author(s):

S.H. Chen ◽

G.E. Seidel

Keyword(s):

Embryonic Development ◽

Intracytoplasmic Sperm Injection ◽

Oocyte Activation

Download Full-text

Increased disruption of sperm plasma membrane at sperm immobilization promotes dissociation of perinuclear theca from sperm chromatin after intracytoplasmic sperm injection in pigs

Reproduction ◽

10.1530/rep.1.0680 ◽

2005 ◽

Vol 130 (6) ◽

pp. 907-916 ◽

Cited By ~ 19

Author(s):

Mika Katayama ◽

Peter Sutovsky ◽

Boh S Yang ◽

Tom Cantley ◽

August Rieke ◽

...

Keyword(s):

Plasma Membrane ◽

Intracytoplasmic Sperm Injection ◽

Membrane Damage ◽

Fertilization Rate ◽

Sperm Chromatin ◽

Eosin Y ◽

Sperm Plasma Membrane ◽

Perinuclear Theca ◽

Sperm Immobilization

The effects of sperm-immobilization methods on decondensation of sperm chromatin and retention of subacrosomal sperm perinuclear theca (SAR-PT) after intracytoplasmic sperm injection (ICSI) were examined in pigs. Sperm membrane damage caused by different immobilization methods by rubbing with a micropipette without piezo pulses (R), or with a low (L) or high (H) intensity of piezo pulses while rubbing, was assessed by the time required for staining of sperm heads with eosin Y solution. The average time for staining of sperm heads immobilized by the R, L or H treatments was 76, 41 or 26 s, respectively. The fertilization rate following ICSI was increased by sperm immobilization by piezo pulses compared with R, but increased intensity of pulses from L to H did not cause further improvements (29, 48 and 47%, respectively). An immunofluorescence study revealed that H immobilization promoted the dissociation of SAR-PT from sperm chromatin compared with L and R, and it increased the frequency of male pronuclear formation in which chromatin appeared uniformly decondensed. Within vitrofertilization (IVF), SAR-PT disassembled coordinately with sperm chromatin decondensation and it was not detectable around male pronuclei. This was different from most of the oocytes after ICSI in which remnants SAR-PT were detected adjacent to male pronuclei. We concluded that increased damage on the sperm plasma membrane at immobilization improved fertilization rates and decondensation of sperm chromatin after ICSI due to the accelerated dissociation of SAR-PT from the sperm nucleus. Also, the behavior of SAR-PT after ICSI was different from that observed in oocytes after IVF.

Download Full-text