Recent technical updates include smart, networked systems and refined models for catheterization.
In both clinical and pre-clinical settings, researchers often need to infuse compounds or withdraw samples of blood or other body fluids. While there is some overlap, notably the infusion pumps, for the most part the devices used with laboratory animals are completely different from those used with humans. In 1999, Jacobson noted in Healing and Smith’s Handbook of Pre-Clinical Continuous Intravenous Infusion,1 “the tethered infusion systems of today look remarkably similar to those of 1970.” However, as we report in this paper, the pace of technological change has increased remarkably over the past few years. While the infusion models from the 1960s through the 1990s have generally held up, the devices used in these models are in the midst of a substantial transformation.
Systems that automate both infusion and sampling can improve accuracy, operational efficiency, and animal welfare. New methods of connecting fluid lines to experimental animals are reducing failure rates and are much easier to use. Finally, new resources for training and collaboration will continue to develop and spread infusion and sampling best practices in the future.
All of these advancements are aligned with, if not driven by, the 3R principles of reduction and refinement.
Researchers in the fields of toxicology and safety pharmacology are the most demanding users of infusion equipment. Running a conventional infusion study with dozens or even hundreds of animals is highly labor intensive. For example, infusion rates must be calculated to adjust the dosage to each animal’s weight, infusion pumps must be programmed individually, pump alarms must be monitored and responded to, and data must be collected and input into laboratory information management systems (LIMS). Each manual step—and there can be tens of thousands of keystrokes to program a study’s worth of conventional pump data—is an opportunity for an error. Furthermore, the extended time technicians must spend in the animal room working with the equipment can stress the animals.
|Product Type||Product Name||Company|
|Infusion Study Automation Software||AVA WIFI™||AVA Biomedical|
|Infusion Study Automation Software||Orchesta™||Instech Solomon|
|Infusion Study Automation Software||Axios™||Strategic Applications|
|Automated Blood Sampler||Culex®||Bioanalytical Systems|
|Automated Blood Sampler||AccuSampler®||CMA Microdialysis|
|Automated Blood Sampler||ABS2™||Instech Solomon|
|Dried Blood Spot Cards||Whatman® FTA® DMPK||GE Healthcare|
Systems that combine smart, networked pumps with centralized control and monitoring software are now available to automate this process (Table 1). A researcher will set up the program for an entire study just once. The system calculates flow rates for each pump using imported weight tables, and then sends the information to each pump
via a wireless or wired network. The central monitoring PC displays alarms, such as occlusions, and can send that information to remote employees via email or text message. It automatically documents events and user interventions,
replacing the vast majority of paper documentation with electronic data.2 These systems have the potential to revolutionize the processes of large scale infusion studies (Figure 1).
Automated systems to withdraw and store blood samples from rats were first developed in the mid 1990s and have already had a significant impact in the departments that
have implemented them. A computer-controlled set of pumps and valves that withdraw precise samples at programmed time points without disturbing the animal replaces the labor-intensive and stress-inducing process of manual blood withdrawals. These systems require researchers to redesign procedures, but nonetheless acceptance has grown steadily over the past decade, particularly in drug metabolism and pharmacokinetics (DMPK) research.As use has become more widespread, the systems have evolved from initial designs to more reliable, easier-to-use second- and third-generation machines. Some of the new systems can sample from mice or large animals in addition to rats (Figure 2).
One of the most exciting developments in blood sampling is the dried blood spot (DBS) technique.3 Vials of 150-500μL of blood which must be refrigerated and spun down are
replaced with a 15μL drop placed on special paper. DBS samples can then be stored at room temperature and shipped in conventional envelopes. In addition to operational efficiency, this technique’s small blood volume
can mean more samples per animal and thereby a reduction in the number animals needed for a given study. This technique is currently available with cards for manual sampling and will be combined with automated sampling in the near future (Figure 3).
In a traditional tethered infusion model, an animal is catheterized in an initial procedure and the proximal end of the catheter is plugged and tucked under the skin while the animal recovers. In a subsequent procedure, the catheter is exteriorized, a harness or jacket is placed around the animal, and the catheter is routed through a spring tether up to a swivel for further connection to an infusion pump outside the animal’s cage. Related models use implanted buttons, with the catheter externalized through the center of the button, or tail cuffs for externalized tail vein catheters.
|Product Type||Product Name||Company|
|Rat Tethers||FastTether2™||AVA Biomedical|
|Rat Tethers||Vascular Access Harness™, Vascular Access Button™||Instech Solomon|
|Rat Tethers||Quick Connect™ infusion system||Strategic Applications|
|Programmable Implantable Pump||iPrecio®||Primetech Corporation|
|Large Animal Port||InLine™ Port, OmegaPort™||Access Technologies|
|Large Animal Port||Cath-In-Cath™||AVA Biomedical|
|Large Animal Port||PortHold™||Instech Solomon|
Several new systems have been designed for rats that refine these models (Table 2). In each, the catheter is attached to a special harness or button at the time of catheterization, eliminating the second procedure and associated recovery time. These new harnesses or buttons feature connectors such that a mating tether can simply be plugged in when the infusion study is to begin. Similarly, the tether can be easily disconnected when the animal needs to be weighed or when the infusion study is complete. Some of these systems are further designed so that the fluid path seals off when not connected, reducing the chance of contamination and infection and preventing retrograde flow that can lead to an occluded catheter.
Most recently, versions of these harnesses are available with connections for two catheters, simplifying procedures such as simultaneous infusion and blood sampling, or bile sampling, where bile can be routed through an external loop in the harness back to the duodenumunder resting conditions, but then is easily diverted for sampling by connecting a mating two-channel tether (Figure 4).
Ideally, an experimental animal would not be tethered, removing the physical restraint and permitting multiple housing. The implantable Alzet® osmotic pump (DURECT Corporation) has long been an option for rodent infusion for those that can adapt their protocols to the fixed flow rates and reservoir volumes that are available from the manufacturer. The recently introduced iPrecio® pump offers a flexible alternative. The 8g pump is small enough for implantation in rats, but includes a miniature peristaltic mechanism and 900μL reservoir that can be refilled through the skin. The pump is programmed prior to implantation and can administer simple or complex flow profiles with rates from 1 to 30μL/hr (Figure 5).
Larger animals can wear a jacket with a pocket for an ambulatory infusion pump. In this case, vascular access is often gained via an implanted port similar to those used in human medicine. Here too there have been advances. While implanted ports can remain patent for years, failures can occur when the animal dislodges the needle from the port.The PortHold™ features a titaniumplate under the septum with holes that are slightly larger than the needles used to access the port. These holes prevent the needle from working it sway out of the port as the animalmoves. The Cath-In-Cath ™ port and the InLine™port prevent needle dislodgements by advancing a percutaneous, through the- needle catheter (instead of a port needle) several centimeters through and past the port septum (Figure 6).
Training and Collaboration Resources
Researchers that are new to animal infusion or sampling now have many resources to draw on. Hands-on training courses in microsurgery are held in specially designed facilities, or instructors can develop a custom course to be held at your site. Online courses are available to users around the world in subjects as specific as rodent catheterization and osmotic pump implantation.
For those that do not have the surgical expertise in-house, most animal vendors offer catheter implantation as a surgical service. Furthermore, these vendors can also install the new harnesses discussed above, reducing the burden on the researcher when the animal arrives to a simple connection of a mating tether.
Experienced researchers also have new resources (Table 3). Discussion groups on internet sites such as LinkedIn are now commonplace. Harlan Laboratories has held several focused “Infusion Seminars” in Europe over the past several years, with participants from pharmaceutical companies, contract research organizations (CROs), academia and industry presenting and sharing ideas. For example, Harlan Laboratories Switzerland presented their efforts to optimize their equipment and procedures, focused primarily on long-term catheter patency, in order to extend their infusion studies out to 13 weeks.4
|Seminars; Training Courses; Publication||Academy of Surgical Research|
|Training Courses, live||Charles River Laboratories|
|Training Courses, live||Columbia University Medical Center Microsurgery Research & Training Laboratory|
|Seminars; Training Courses, live||Harlan Laboratories|
|Seminars; Publication||Infusion Technology Organisation|
|Training Courses, live||René Remie Surgical Skills Centre|
|Training Courses, online and live||Veterinary Bioscience Institute|
Taking this concept one step further, in 2009 a group of researchers in Europe working in different, and sometimes competing, companies formed the Infusion Technology Organisation. Its goals are broad: “to provide a forum by which information and data generated from infusion technology can be shared on a global basis.”
These advances in equipment and techniques are already producing results. For example, Dr. Russell Bialecki at AstraZeneca has combined infusion, automated blood sampling, radio telemetry (measuring parameters including blood pressure, heart rate, temperature, and ECGor EEG), and collection ofmetabolicwaste.With this “integrative pharmacology” model they collect data in a single experiment that would traditionally require several. In addition to the obvious benefits of a reduction in the number of animals and the time and expense to conduct separate studies, often the availability of the simultaneously-collected data can help researchers understand variability that would have otherwise been interpreted as noise.5
Researchers that are able to achieve long-term catheter patency not only have the option to run longer studies, but also to reuse animals after a wash-out period. Refinement methods such as remote control of infusion pumps, video assessment, and perhaps even robotic removal or introduction of fluids from the animal room can further reduce animal stress while streamlining research processes.
As researchers continue to make individual improvements and then share them in the various forums for collaboration—prompting manufacturers to develop new products in the process—the devices and techniques used in laboratory animal infusion and sampling will continue to advance in the future.
- Healing, G. and Smith, D. Handbook of Pre-clinical Continuous Intravenous Infusion. Taylor & Francis, 2000. ISBN 978- 0748408672.
- Haas R., Jacobson A., Sommers, J., Pawl S., Agate J., Brooks A., Alexander A. “Cooperative development of an automated syringe infusion pump for preclinical toxicology.” Poster at 2009 Society of Toxicology Meeting.
- N. Spooner, R. Lad, M. Barfield (2009) Anal. Chem. 81 1557-1563 http://www.ncbi.nlm.nih gov/pubmed/19154107
- Kaiser, S. “Continuous Infusion Toxicity Studies in Rats: Experiences and Developments at Harlan Laboratories Switzerland.” Presentation at Harlan Infusion Seminar, Paris France, 11 May 2010.
- Kamendi, H.W., Brott D.A., Chen, Y., Litwin, D.C., Lengel, D.J., Fonck, C., Bui, K.H., Gorko, M.A. and Bialecki, R.A. “Combining Radio Telemetry and Automated Blood Sampling: A Novel Approach for Integrative Pharmacology and Toxicology Studies.” Journal of Pharmacological and Toxicological Methods (2010), doi: 10.1016/ j.vascn.2010.04.014