Research: Artificial Insemination Technology

Increased fundamental knowledge of the cellular and molecular
mechanisms regulating oviductal sperm transport, selection and storage
will provide logical, science-based approaches toward the development
of novel semen diluent formulation and semen storage techniques.

Increased fundamental knowledge of the cellular and molecular mechanisms regulating oviductal sperm transport, selection and storage will provide logical, science-based approaches toward the development of novel semen diluent formulation and semen storage techniques.

Recently, we have examined not only on how sperm survive in the uterovaginal junction (UVJ) sperm-storage tubules (SST), but the role of highly specialized newly discovered cells in the vaginal epithelium.  These cells, which have been tentatively identified as non-neuronal endocrine cells (NEC) may influence sperm metabolism and motility in the vagina through their secretory activities.  The NEC are found in the intestinal epithelium of mammals and aves, and are more commonly referred to as enterochromaffin cells. Their major secretory product is the neurotransmitter serotonin.

Here we will briefly review some of our findings and speculate on their possible implications to artificial insemination (AI) technology.

In poultry, the hen’s oviduct is a single long, convoluted tube that functions in the assembly of a hard shell egg. The oviduct also is the site of fertilization and early embryo development.

The importance of the SST cannot be underestimated.  Maximum, sustained fertility will only be attained in the turkey hen when the SSTs are filled with sperm.  In the turkey, this is best accomplished with inseminations just prior to the onset of egg production.  Initial inseminations after the onset of egg production will result in a less than optimal number of sperm in the SST and consequently, lower fertility.  The mechanisms responsible for prolonged sperm survival within the SST have yet to be fully determined.  The events that appear to be responsible for successful sperm storage and release in the SST are listed in the coloured box.

The vagina is the terminal segment of the oviduct and is most frequently  thought of as the site of semen deposition after AI or more simply as a conduit for the egg mass passing from the uterus through the cloaca at egg laying. There are also less appreciated functions of the vagina that are vitally important to the reproduction process. One such function is the process of sperm selection.  Of the 250-300 million sperm inseminated, fewer than 5 million are ever stored in the SST at one time.  This is a very intense selection process with presumably, only the most fit sperm selected to be transported to the SST. 

Unfortunately, we know little of the cellular and molecular controls regulating sperm selection.  This sperm selection process can also be viewed as sperm competition, that is, competition between sperm of the same male or sperm between males to gain access to the SST and ultimately fertilize the ova.  

We know that those sperm exhibiting the greatest ability to penetrate a viscous medium in vitro, a measure of sperm mobility, are the most likely sperm to reach and fertilize the ova.  By selecting toms based on their in vitro assessment for sperm mobility, fertility in the turkey hen can be improved. From this, it is apparent that sperm competition is, in part, influenced by sperm characteristics (i.e. phenotype).  

How does the vagina impact sperm competition?  Can the vagina influence which sperm reach the SST?  Histologically, the vagina is unique when compared to the other segments of the oviduct. While the vagina’s mucosal surface is lined with a pseudostratified columnar epithelium consisting of predominately ciliated cells, there are no subepithelial tubular glands. 

In addition, in opportunistic sections of vaginal mucosa, a cell in the surface epithelium that morphologically does not resemble either a ciliated or secretory cell is occasionally observed.  These cells, the presumptive NECs, have a broad basal region and more tapered apical region, resembling a bowling pin.

While we have yet to confirm that these are NEC by histochemical procedures specific for some gut enterochromaffin cells, serotonin-positive cells in the vaginal surface but not the SST epithelia have been observed.   

The presence of serotonin-producing cells in the vaginal epithelium may have significant impact on vaginal sperm selection and transport to the SST, hence sperm competition.  In the gut, the enterochromaffin cells sense, either through mechanical or chemical stimulation, the presence of luminal material, and release serotonin.  The serotonin released from the enterochromaffin cells reach sensory neurons in the lamina propria and initiate a locally controlled peristaltic contraction. 

Interestingly, in other animal and cell systems, serotonin has been shown to increase the tail-beat frequency in sperm as well as the cilia beat frequency in ciliated epithelial cells.  So hypothetically in the turkey vagina, inseminated sperm, some of which expressing a unique molecular or metabolic characteristic (the subpopulation of ‘fit’ sperm), trigger the NEC and the local release of serotonin or another serotonin-like substance.  The cilia on the ciliated cells increase in beat frequency as do the sperm tails, and peristaltic-like contractions ensue, all of which potentiating sperm transport to the UVJ-SST. 

If this hypothesis proves to be true, the implications could eventually be very significant to those of us seeking to
reduce the number of sperm inseminated per AI while increasing the interval between successive inseminations.  Identifying the signals or characteristics associated with sperm that activate the serotonin-cascade, or more simply, adding serotonin to the AI dose or diluent may result in increased numbers of fit sperm reaching the SST throughout the egg laying cycle.  Increased number translates into increased hen fertility. n
This report was presented at the Midwest Poultry Federation Convention.  For further information, contact Murray R. Bakst, e-mail: murray@anri.barc.usda.gov.