ABSTRACT
Interest in ion channels as drug targets for contraception has grown with the realization that certain ion channel subunits are located exclusively in sperm. Selective knockdown of ion channel subunits can lead to infertility without ill effects, and selective inhibitors and/or openers of these ion channels could interfere with sperm function. In this study, in vivo electroporation (EP) and rete testis microinjection-mediated plasmid DNA were adopted to silence CatSper2 expression, which is essential in sperm hyperactivation. The results showed that high transfection efficiency and expression were achieved by plasmid DNA that was directly injected into the rete testis. As a result of the expression of CatSper2 being blocked, the treatment group showed significantly lower (P<0.05) hyperactivation rate, fertilization rate in vitro, migration motility in viscoelastic solution and intracellular Ca(2+) peak. The low hyperactivation and fertilization rates lasted for 60 days. Meanwhile, analysis of the sperm survival rate and testis histology indicated that in vivo EP had no significant effect on the function of the testis, spermatogenesis or sperm activity. The present study demonstrated that it was feasible to achieve male contraception by silencing the expression of CatSper2, the key protein involved in sperm hyperactivation.
Subject(s)
Calcium Channels/physiology , Fertility/physiology , Fertilization/physiology , Gene Knockdown Techniques , Seminal Plasma Proteins/physiology , Animals , Calcium Channels/genetics , Cell Survival/drug effects , Male , Models, Animal , RNA, Small Interfering/pharmacology , Rats , Rats, Sprague-Dawley , Seminal Plasma Proteins/genetics , Signal Transduction/physiology , Sperm Motility/physiology , Testis/drug effectsABSTRACT
The objective of this study was to investigate the variation of HSP70 mRNA level in dairy cows and relationships of its closely linked microsatellite loci with heat tolerance traits. Blood samples were collected from ten healthy Holstein cows with the same age and milking stage at different temperatures-humid-index (THI) (86.2, high temperature; 70.9, critical high temperature, and 56.8, optimum temperature). The mRNA levels of HSP70 of lymphocytes in peripheral blood were analyzed using real-time RT-PCR. The mRNA level of HSP70 was increased with the THI; the mRNA level of HSP70 at high temperature was higher than others (P<0.01). This indicated that the bovine HSP70 gene may act as a potential can-didate gene for response to heat shock. Genetic variation of three microsatellite loci BMS468, BM1258, and BM1815, which were closely linked to HSP70 gene on chromosome 23, was analyzed in 160 Holstein cows with non-denaturing poly-acrylamide gel electrophoresis. The association between these microsatellite loci and heat tolerance traits were analyzed by least square linear model. The results showed that 134 bp/128 bp at BMS468 and 186 bp/148 bp at BM1815 were the most favorable genotypes for HTC, red cell potassium, and decrement rate of milk yield in high temperature (P<0.05); 101 bp/99 bp at BM1258 was the most favorable genotype for decrement rate of milk yield in high temperature (P<0.05).
Subject(s)
HSP70 Heat-Shock Proteins/genetics , Hot Temperature , Microsatellite Repeats/genetics , Milk/metabolism , RNA, Messenger/physiology , Animals , Cattle , Female , Lymphocytes/metabolism , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction , TemperatureABSTRACT
OBJECTIVE: To evaluate the role of sperm kinematic parameters in the hyperactivation of Guinea pig spermatozoa, and to confirm the index of their hyperactivated motility. METHODS: Computer-aided sperm analysis (CASA) was used to describe the kinesis parameters of the Guinea pig spermatozoa incubated for 1, 3, 5 and 7 hours. RESULTS: The curvilinear velocity, average path velocity and amplitude of lateral head displacement were increased with time and reached the peak at 5 hours, while the straight linear velocity, linearity, straightness and beat cross frequency were gradually decreased with time and hit the bottom at 5 hours. CONCLUSION: The sperm movement pattern changes greatly before hyperactivation during the capacitation of Guinea pig spermatozoa.
