|T H E N I H C A T A L Y S T||N O V E M B E R D E C E M B E R 1999|
|H O T||M E T H O D S|
Tory Hampshire, DVM, NINDS
Mouse Tips: A Three-Part Series
PART 3. THE ROUTES TO RIGHT DOSING
Parts 1 and 2 "inside the mouse hospital" addressed old and new ideas for surgery and perioperative support of the mouse and rat. Part 3 presents some available products and techniques that may make medication of the mouse less labor-intensive and more efficient.
Some General Tips
Dosing and achieving steady-state blood concentrations are challenging in a tiny patient with a high metabolic rate whose surface-to-volume ratio is roughly ten times that of humans.
When an oral route of administration is selected, its a good idea to measure pooled blood samples via retro-orbital bleeds (under anesthesia) several times over the course of 24 hours to make certain that steady-state blood concentrations are achieved. Often, this step is left out of study design. A volume per mouse of 100 mL is acceptable. Alternatively, mice can be terminally bled, serially, to construct a pharmokinetic curve.
A common myth is that mice and rats are resistant to infection, but in reality even oral dosing entails a risk of infection in genetically or pharmacologically immunosuppressed mice. Always pay attention to osmolality, pH, and sterility of preparation in dosing immune-compromised hosts like SCID or nude mice. And remember that although a healthy immune-competent mouse may have an LD50 for infection somewhat higher than that of a person, a dog, or a cat, it does have an LD50. For intravenous, intramuscular (IM), or intraperitoneal injections, continue to observe precautions in preparing drug solutions or fluids.
Instech Companys single-channel infusion system uses the Harvard Apparatus infusion pump, stainless steel single-channel swivel (25 gauge), 3.5" counterbalanced lever arm, covance infusion harness for mice, and an 8.5" clear animal container with feeder and water bottle.
The Infu.Disk by Med.e.cell (San Diego, 619-552-0781; or website ) can be mounted to a swivel arm and serves as a lower-cost alternative to the Harvard system for infusion.This company makes five different 10-mL discs delivering a range of 0.024.0 mL/h.
Most scientists use gavage needles to deliver substances via the oral route. Gavage needles are readily available from almost any scientific supply source. Stomach capacity is generally 510 mL/kg or about 0.2 mL at a time for a mouse. Some mice will willingly drink off the end of a gavage needle if the substance is highly palatable. Gavage needles should be checked for rough edges as the esophagus in mice is very delicate.
The Jell-O recipe mentioned in our first article is also highly palatable and may also serve for noncritical dosing regimes. Keeping in mind that mice typically revisit food throughout the day, it might be wisest when using Jell-O as a medication vehicle to place the total desired daily dose in one Jell-O cube of known volume. The same considerations should guide the process of adding medications to water and pelleted feed, a popular way to administer drugs orally. Again, measure pooled aliquots of serum before arriving at conclusions about drug efficacy. We have had good luck with a source called National Medical Services (Willow Grove, Penn., at 215-657-4900 or e-mail, which runs a large number of bioanalyses for a broad range of compounds and will also consult on special problems.
Because of the large space between skin and subcutaneous tissues, the subcutaneous route is very attractive for chronic medication of rodents. Generally, it is very easy to restrain mice and to deliver up to 5.0 mL in this space. For optimal absorption, drugs should be hypoosmolar (less than 300 milliosmoles). Necrosis at the site of entry is a major side effect with highly acid or base substances, but sterility is not as much of a problem with subcutaneous administration as it is with the intramuscular, intraperitoneal, or intravascular routes. Immunosuppressed mice, SCID mice, and nude mice should not receive anything that has not been carefully sterilized.
Slow- and continuous-release options have also become available in transcutaneous and subcutaneous rodent delivery systems.
Several catheter delivery systems are also available for rodents. Alzet osmotic pump models allow continuous delivery of agents at controlled rates when placed subcutaneously or intraperitoneally. If you want targeted delivery of a drug to an area remote from the site of implantation, you can attach a catheter to the pump. Alzet pumps have been used in gene therapy experiments. The pumps come in several sizes with different volume reservoirs. For more information, visit the web site or call 800-227-9953 or e-mail.
The ESOX implantable pump is also an option. This system, with its refillable reservoir, enables truly long-term drug delivery. (Contact Access Technologies or 847-674-7131.)
Innovative Research of America (Sarasota, FL, 800-421-8171) has come out with a time-release matrix-driven biomedical delivery system in a pelleted form that easily can be placed in a subcutaneous pocket. They list nearly all compounds of drugs and do special formulations as well.
The advent of pain medications in topical gels and creams is here. Systemic absorption of active drugs after topical application of ketoprofen (a nonsteroidal anti-inflammatory), nitroglycerin, motion-sickness drugs, and the like has led to a growth industry of creating and evaluating penetrating emulsions. Organogel, a lecithin-based matrix, has been used by our consulting pharmacist (Foers pharmacy, Bethesda, MD, 301-657-3500) to compound 40 percent (38 percent stronger) lidocaine cream for studies confounded by narcotics and nonsteroidals. We apply this along the incision line of rodent patients. It may represent another avenue for innovative stress-free application of drugs.
As we mentioned in our second article, with lots of practice, it is possible to cannulate mouse jugular veins. Its definitely possible to purchase mice and rats already instrumented with intravascular accessfrom Taconic Farms (through the Veterinary Resources Program procurement, 301-496-3575)or put them in yourself. In mice, if you are successful with this route, several intravenous delivery solutions are possible. Harvard makes a terrific mouse infusion system, with a swivel that has low rotational friction. This system, with all attachments, runs around $3,500.00 and is sold through Instech (5209 Militia Hill Road, Plymouth Meeting PA, at 610-941-0132; fax: 610-941-0134).
IM and Intraperitoneal Delivery
We discourage the use of IM injections. Necrosis and pain at the injection site are not uncommon outcomes due to the small hindlimb muscle mass of rodents. If you must use the IM route, you should learn the location of the sciatic nerve and limit the volume delivered to 0.050.1 mL/site in mice and.0.10.3 mL/site in rats. The lumbar muscles are also good IM injection sites. Due to the small volume per site, one drug dose generally must be delivered in several places.
Intraperitoneal delivery is still overused in rodent studies and may carry with it numerous concerns, such as contamination, splenic trauma, serosal hemorrhage, and other untoward effects, and should be performed thoughtfully. Drug delivery by convection or viral-vector delivery to the spleen or liver is best performed under direct visualization of these structures. Drug delivery is possible but must follow general guidelines for sterility, pH, and osmotic compat-ability with mammalian systems.
Attention to such detail and careful practice with NIHs tiny research subjects will surely pay off with better data.
|Disclaimer: Mention of specific products in this article does not constitute an endorsement of those products, nor does it signify that other similar products are less desirable.|
After 11 years, Tory Hampshire is leaving NIH in December to start her own businessAdvanced Veterinary Applicationsin Bethesda. Shell be focusing on refinements to animal care and veterinary technician team building.Ed.
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