By Kimber Liedl
Watching a common marmoset mother carrying around two tiny infants, one would expect that both babies would be her own. But what if one infant turned out to be both aunt and half-sister to the other?

“Marmoset families are more complex than we used to think,” says Wendy Saltzman, Ph.D., co-principal investigator with David Abbott, Ph.D., on studies of reproduction and family dynamics in the common marmoset (Callithrix jacchus). Center studies on reproductive suppression and hierarchical change in relation to “stepfather” introductions are shedding new light on previous theories of marmoset family structure.

“The traditional belief has been that there is always a single dominant breeding female in any social group,” Saltzman said. “But increasing evidence from both the lab and the field shows this is not always the case.”

Reproductive failure in subordinate females has been previously attributed to physiological and behavioral suppression imposed by their dominant mothers. By 1993, however, Saltzman also began to question the role of males in reproductive suppression. She hypothesized that inbreeding avoidance may also play an important role in reproductive failure: Daughters living with their natal families might not breed because the only available males are close relatives, that is, their fathers and brothers.

To test her hypothesis, Saltzman began a pilot project to replace the breeding males in marmoset family groupings with unrelated males, or “stepfathers.” She expected that the daughters would be able to escape from reproductive suppression and begin breeding, even with the dominant females still present.  She theorized that the scenario might even lead to simultaneous breeding by both mothers and daughters.

The pilot project was successful. “Some of the ovulating daughters copulated and became pregnant in response to the introduction of the stepfathers,” Saltz-man said. By contrast, in intact, or undisturbed families, about half the daughters may ovulate, but typi-cally none breed, as they have no sexual interaction with their father or brothers.

The resulting, full research project funded by the National Science Foundation began in early 1998, with 11 “stepfather” families and seven control families. Saltzman and her collaborators collected behavioral data, blood and urine samples from daughters and mothers. When there was a possible pregnancy, David Abbott used ultrasound scans to confirm and monitor the pregnancy every four weeks. Behavioral tests such as temporary separations and reunions helped researchers examine the dominance relationship between mothers and daughters.

Of the 11 stepfather families in the study, eight daughters had offspring. In contrast, none of the daughters in the control families copulated or conceived. Furthermore, ovulating daughters—whether in stepfather or control families—were not suboordinate to their mothers, whereas anovulatory daughters were suboordinate. Thus, Saltzman’s findings point to at least two components of reproductive failure in marmoset daughters: First, ovulation suppression is associated with a daughter’s relationship to her mother. Second, inhibition of sexual behavior is closely tied to lack of unrelated males in the social group. “Therefore, in order to breed, daughters must be both insubordinate to their mothers and have access to an unrelated male, “ she explained.

In addition to observing that captive common marmoset daughters can escape from reproductive suppression when an unrelated male is present, Saltzman also noticed some surprising family dynamics. For example, even though there could be several adult-aged daughters in a family, only one daughter at a time would breed with an introduced stepfather. “This was usually the oldest and highest ranking daughter, but we saw cases where a younger daughter outranked her older sister and began to breed,” Saltzman said. In addition, two breeding females—mother and daughter—would carry and nurse each other’s infants. However, aggression also increased and threatened infant survival.

Furthermore, Saltzman said, if an older, breeding daughter was later removed from the family, the remaining younger daughter next in rank would begin breeding if the stepfather was still present.

When a mother and daughter both bred with a stepfather, sometimes one female would become very protective of her infants, shielding them from other family members. Yet the introduction of the new male did not create direct problems for infant survival. Unlike many primate species, Saltzman explained, marmosets are cooperative breeders, and stepfathers do not try to kill dependent offspring. Males are often very attracted to parenting infants even if they are not their own offspring.

Saltzman believes such research might help explain situations in the field where two breeding females are observed in a group or where there is reduced infant survival. New genetic data from wild groups of common marmosets in Brazil are consistent with her results in captivity. Studies by Nievergelt et al, 2000, revealed that, in wild groups with two breeding females, those females are closely related to each other.

“They are probably mothers and daughters, or they are sisters,” Saltzman said. “They are also related to all other group members except the breeding male. This suggests that males may regularly transfer between groups in the wild.”

Saltzman’s research could aid in the management of captive common marmoset colonies at research centers and zoos. She explains: “Although it may be tempting to look at these alternative families as a possible method to produce more offspring per family, the traditional, one-breeding female group appears to minimize overall aggression and maximize infant survival.” For instance, in many captive marmoset colonies, she said, if a breeding male dies, he is routinely replaced by an unrelated male. “However, we now know that if a daughter is present in the family, even if she is not yet mature, this can lead to problems later on.”

Dr. Wendy Saltzman has been conducting this research to lead to a better understanding of marmoset breeding behavior in the wild, and to more knowledgeable management of captive marmoset colonies. She will continue as an active affiliate scientist with research assistants at the Primate Center after she moves to California this September. She has accepted a position as an adjunct assistant professor of biology at the University of California at Riverside (http://cnas.ucr.edu/~bio/faculty/Saltzman.html).

Reference:
Saltzman W, Schultz-Darken NJ, Severin JM, Abbott DH. 1997. Escape from social suppression of sexual behavior and of ovulation in female common marmosets. Ann N Y Acad Sci. Jan 15;807:567-70.
 

Collaborators refine research on male contraceptive

David Abbott, Ph.D., Pam Tannenbaum, Ph.D.,  and undergraduate directed study student Melissa Gannage are collaborating with Vichai Reutrakul, Ph.D., visiting scientist from Mahidol University in Thailand, to study the effects of a natural male contraceptive in common marmosets.

Their grants are through the UW Asian Partnership Initiative and Contraceptive Development Program (CONRAD; a component of US AID).

As one of about 20 new UW research collaborations started through this initiative, Dr. Reutrakul is working with the WRPRC team to isolate a chemical called triptolide from an Asian plant species. The contraceptive effects of triptolide have been known since the 1970s; yet this is the first controlled study in a primate species. 

Drs. Abbott, Tannenbaum and Reutrakul are concluding a three-year successful project on triptolide as a contraceptive in male marmosets. They expect to publish on their results in Fall 2001.
 

Just published

Following is a sample listing of recent publications involving WRPRC resources. Abstracts for these and other center publications can be viewed at the National Library of Medicine’s PubMed Web site. 

Allen TM, Mothe BR, Sidney J, Jing P, Dzuris JL, Liebl ME, Vogel TU, O'Connor DH, Wang X, Wussow MC, Thomson JA, Altman JD, Watkins DI, Sette A. CD8(+) lymphocytes from simian immunodeficiency virus-infected rhesus macaques recognize 14 different epitopes bound by the major histocompatibility complex class I molecule mamu-A*01: implications for vaccine design and testing. J Virol 2001 Jan;75(2):738-49.

Cho YG, Ramer J, Rivailler P, Quink C, Garber RL, Beier DR, Wang F. An Epstein-Barr-related herpesvirus from marmoset lymphomas. Proc Natl Acad Sci U S A 2001 Jan 30;98(3):1224-1229.

Dejesus OT, Endres CJ, Shelton SE, Nickles RJ, Holden JE. 2001. Noninvasive assessment of aromatic L-amino acid decarboxylase activity in aging rhesus monkey brain in vivo.
Synapse. Jan;39(1):58-63.

Kemnitz JW, Roecker EB, Haffa AL, Pinheiro J, Kurzman I, Ramsey JJ, MacEwen EG. Serum dehydroepiandrosterone sulfate concentrations across the life span of laboratory-housed rhesus monkeys. J Med Primatol. 2000. Oct;29(5):330-337.

Ramsey JJ, Colman RJ, Binkley NC, Christensen JD, Gresl TA, Kemnitz JW, Weindruch R. Dietary restriction and aging in rhesus monkeys: the University of Wisconsin study. Exp Gerontol. 2000. Dec 15;35(9-10):1131-1149.

Schramm, RD, and Paprocki, AM. 2000.  Birth of rhesus monkey infant after transfer of embryos derived from in vitro matured oocytes: short communication. 2000. Hum. Reprod. Nov;15(11):2411-2414.

Sidney J, Dzuris JL, Newman MJ, Johnson RP, Amitinder K, Walker CM, Appella E, Mothe B, Watkins DI, Sette A. Definition of the mamu A*01 peptide binding specificity: application to the identification of wild-type and optimized ligands from simian immunodeficiency virus regulatory proteins. J Immunol. 2000 Dec 1;165(11):6387-99.

Terasawa E, Fernandez DL. Neurobiological Mechanisms of the Onset of Puberty in Primates. Endocr Rev. 2001. Feb 1;22(1):111-151.

Torre P. Lasky RE. Fowler CG. Aging and middle ear function in rhesus monkeys (Macaca mulatta). Audiology. 39(6):300-304, 2000 Nov-Dec.

VanderHorst VGJM, Terasawa E, Ralston HJ, Holstege G. 2000. Monosynaptic projections from the nucleus retroambiguus to motoneurons supplying the abdominal wall, axial, hindlimb, and pelvic floor muscles in the female rhesus monkey. Journal of Comparative Neurology. Aug 21;424(2):233-250.
 

Primate Center hosts workshop on aging May 31-June 1

The Wisconsin Regional Primate Research Center and the National Institute on Aging are hosting a workshop entitled, “Nonhuman primate models of aging: Evaluating their current status and future potential,” May 31 through June 1, 2001, at the Monona Terrace Convention Center, Madison, Wisconsin.

The workshop precedes the 30th Annual Meeting of the American Aging Association at the convention center June 2-4. Registration is Friday, June 1, 5 to 6 p.m. Immediately following registration is a welcome reception and special poster session on Nonhuman Primate Models of Aging and Caloric Restriction in Nonhuman Primates.

For information on both the workshop and the 30th Annual meeting of the American Aging Association, please contact:

Donna Cini, American Aging Association
The Sally Balin Medical Center, 110 Chesley Drive, Media, PA  19063
Phone:  610-627-2626
Fax:  610-565-9747
Website:  http://www.americanaging.org .

Local contact:
Edi Chan, Wisconsin Regional Primate Research Center
1220 Capitol Court, Madison, WI  53715-1299
Phone:  (608) 263-3500
Fax:  (608) 263-4031
E-mail:  chan@primate.wisc.edu.
 
 

Gleanings

New grants
WRRPC affiliate Ted Goodfriend, M.D., Depts. of Pharmacology and Medicine, has been awarded a three-year grant by the American Heart Association to study fatty acid metabolites that may contribute to the hypertension accompanying obesity in rhesus monkeys.

Jon Ramsey, Ph.D., has received a one-year grant from Pfizer Inc., entitled, “Regulation of food intake in rhesus monkeys.” To investigate the neuroendocrine regulation of food intake in rhesus monkeys, he plans to test the effects of several newly discovered neuropeptides on food intake, energy expenditure and body composition.

Mary Schneider, Ph.D., Harlow Center for Biological Psychology and Department of Kinesiology, has received a five-year, $2 million grant from the NIH?National Institute on Alcohol Abuse and Alcoholism. The five-year grant is titled, “Moderate level prenatal alcohol exposure in primates.”

Dee Schramm, Ph.D., received a $3 million five-year NIH grant for production of MHC-defined and genetically identical pairs of MHC-defined monkeys using assisted reproductive technology.

Charles Snowdon, Ph.D., UW Psychology Department, has a new, $1.45 million NIMH grant entitled “Evolution and Development of Primate Speech Analogues.” The grant is funded through April 2006.

David Watkins, Ph.D., has two new NIH grants, administered through the UW Department of Pathology. A three-year grant is entitled “MHC Typing of Macaques used in AIDS research.” A four-year grant is entitled “A novel, logical approach to HIV vaccine development.”

Richard Weindruch, Ph.D., and Tomas Prolla, Ph.D., have received an NIA grant, “Gene Expression Profiling, Oxidative Stress and Aging,” The grant is funded from March 1, 2001, to Feb. 28, 2006.

Honors
Richard Weindruch, Ph.D., has received the annual Glenn Foundation Award from the Gerontological Society of America (GSA). He is honored for his contributions to the field of aging, especially to the new understanding of the relationship among caloric intake, oxidative stress and aging. Weindruch’s recent groundbreaking research with colleagues Tomas Prolla, Ph.D., and Cheol-Koo Lee, M.S., using oligonucleotide microarrays strengthens the idea that caloric restriction may slow aging by reducing oxidative damage to tissues.

Appointments, elections
Wendy Saltzman, Ph.D., has accepted an adjunct assistant professorship at the University of California, Riverside, beginning in the 2001-02 school year. She will remain an active affiliate of the Primate Center as she has a continuing grant here to study adrenocortical suppression in common marmosets. Dr. Saltzman and her husband, Ted Garland, Ph.D., UW Zoology Dept., will be working in the Department of Biology at Riverside.

Ei Terasawa, Ph.D., was appointed to the editorial boards of Endocrinology, official journal of the Endocrine Society, and Experimental Biology and Medicine, official journal of the Society for Experimental Biology and Medicine. Both appointments run from  January 2001 to December 2003.

Richard Weindruch, Ph.D., was recently appointed a member of the external advisory board for the University of Southern California’s Department of Cell and Neurobiology, on the NIH Program Project, “Dopaminergic and Basal Ganglia Plasticity and Aging.” Dr. T.H. McNeill is the principal investigator.

Joe Kemnitz, Ph.D., has joined the external advisory board for the University of Texas Health Sciences Center?San Antonio, Department of Physiology. He is advising on the NIH program project, “Nutritional Probe of the Aging Process.” Dr. Arlan Richardson is the principal investigator.

Promotions
Stacey Maves, Animal Services, received her Laboratory Animal Training (LAT) certification from the American Association of Laboratory Animal Science (AALAS) on Dec. 5.

In the news
Joe Kemnitz, Ph.D., was interviewed in November by John Travis of Science News, on dietary restriction and cancer) and by Joanie Stewart of CBS Healthwatch, on nutrition and aging.

Karen Strier, Ph.D., was featured Oct. 25 in Wisconsin Week’s, “Lurking with muriquis—A glimpse of what life could be like for us.” View on line at http://www.news.wisc.edu/view.html?id=5464.

David Watkins, Ph.D., Todd Allen, Ph.D., and David O’Connor, Ph.D., were interviewed by Linda Berris for “MHC typing of monkeys enables cutting-edge AIDS research.” The Fall 2000 NCRR Reporter article appears at http://www.ncrr.nih.gov/newspub/oct00rpt/Mhctyp.htm.

New staff
Sarah Fuenger, B.S., associate research specialist, Jan. 1 (Watkins Lab).
Matt Hoffman M.A., Internet Services librarian, Jan. 1 (Library).
Thomas W. Lynch, B.S., Data Management and Imaging Unit manager, Nov. 1.
Cai-Ping Ren, Ph.D., research associate, Oct. 5 (Thomson lab).
Trevor Richter, Ph.D., research associate/postdoctoral research fellow, Dec. 4 (Terasawa lab).
Wendy Schliewe, B.S., associate research specialist, Feb. 26. (Ramsey lab).
Jim White, B.A.,WRPRC Colony Manager, Feb. 5

Departures
Melissa Browne, B.A., Kemnitz lab, Reproduction Research Services, Library, March 4.
Lisa Knowles, M.S., Reproduction Research Services, Oct. 27.
Vivienne Marshall, Ph.D., Reproduction Research Services, Thomson lab, Jan. 31.
Marian Piekarczyk, Ph.D., Watkins lab, Jan. 31.

WRPRC Research Highlight—Monkeys reveal metabolic mysteries

Heavy monkeys, lean monkeys, monkeys doing backflips, monkeys sitting calmly. One by one, assistant scientist Jon Ramsey, Ph.D., coaxes each animal out of its cage, into a transport cage, and into a special plexiglass enclosure where they can be easily photographed for research and publication needs.

Ramsey spends most workdays among his monkeys and can tell them all apart, even though many are among the 79 animals involved in the Primate Center’s largest research project to date. This is the aging and caloric restriction study, led by Richard Weindruch, Ph.D. The study has been funded by the National Institute on Aging for the past 12 years.

Ramsey, who came to the Primate Center as a post-doctoral trainee working with Joseph Kemnitz, Ph.D., in 1995, now holds the reins to three projects investigating aging and the neuroendocrine regulation of food intake. He is looking at energy expenditure, or oxygen consumption, at the whole animal, organ and cellular levels. Ramsey’s central hypothesis is that caloric restriction without malnutrition increases maximum lifespan by decreasing oxygen consumption and mitochondrial reactive oxygen production.

“Studying energy expenditure changes at different levels can expand our knowledge of how to better target treatments for various diseases related to aging, Ramsey says.

Ramsey has recently discovered that caloric restriction causes an initial decrease in energy expenditure, or metabolic rate. This decrease, however, disappears with time and is not uniform across all organs.

During the first year of the study, Ramsey expected that the calorie restricted monkeys would expend less energy than the controls. They were eating 30 percent fewer calories and consuming less oxygen. After 10 years, however, the restricted monkeys, although they were markedly thinner, were turning out to be no less active than the controls. Ramsey analyzed his data and determined that, after the first year and over time, all the restricted monkeys had gone back to expending the same amounts of energy as the controls. Measurements were expressed as energy expenditure per unit of lean mass.

Then, in another twist, as study hit the 10-year study mark, the restricted monkeys once again started showing a decrease in energy expenditure. This intrigued Ramsey, who thought, “How can we better measure what we’ve been observing at the whole animal level?” He pondered that no one knew the effects of long-term calorie restriction beyond looking at whole animal metabolic rates, and that there would be many different ways to measure it.”Organs, tissues and cells all have different rates of oxygen consumption,” he pondered. “So we needed to take a much closer, less superficial look.”

Scientists have known for a long time that animals with low metabolic rates, including humans, tend to live longer. They also know that caloric restriction decreases metabolic rate, at least initially. Now, Ramsey is able to measure for the first time how this decrease might have lasting effects on animals through the impact on their individual organs.

Ramsey has come up with a novel way of “looking inside” a monkey’s organs. Lightweight cloth mesh jackets temporarily hold catheters placed into various organs—skeletal muscle, brain, liver. Ramsey can then measure each organ’s individual oxygen needs and consumption. Over time, these measurements will give him a clearer picture of how dietary restriction affects individual organs in both the short and long term, and, subsequently, the whole animal as it ages.

“The internal organs are responsible for greater than 60 percent of resting oxygen consumption,” Ramsey explains. “The mass of several internal organs, especially the liver and gastrointestinal tract, is rapidly decreased at the onset of dietary restriction. Without accounting for differences in the effects of dietary restriction on internal organs, it is impossible to determine whether dietary restriction causes a uniform decrease in oxygen consumption in the whole animal over time.”

To delve further into the mysteries of metabolic changes, Ramsey is using rats as a precursor to nonhuman primate studies to investigate changes in energy expenditure at the cellular level. The burning question to answer here is whether there are smaller, cellular changes in oxygen consumption that may be masked by larger changes in organs shrinking at different rates. Ramsey is examining a process called mitochondrial proton leak, which is a major contributor to cellular energy expenditure.* His initial studies show that mitochondrial proton leak decreases with calorie restriction.

“To me, this is the most interesting process and may be the most changed with calorie restriction and the most critical to the aging process,” Ramsey says. “Proton leak is responsible for 20-25 percent of whole animal energy expenditure—that’s using up the oxygen from every fourth or fifth breath you take.”

Ramsey aims to learn how this mechanism is affected by calorie restriction, first in rats, then in primates, then, ultimately in people. He points out that proton leak is highest in skeletal muscles and brain tissue. “This process could be playing a role in people’s susceptibility to sarcopenia or Alzheimer’s Disease,” he ventures.

Ramsey’s body of work is also adding weight to the free radical theory of aging. Oxygen free radicals are another culprit that may be damaging cells, leading to organ deterioration and visible signs of aging. Burning added calories consumes more oxygen, which may be accompanied by the production of excess free radicals. These highly reactive metabolic byproducts bounce around and wear down cells, tissues and organs. A calorie-restricted monkey consumes less oxygen and may produce fewer free radicals.

Finally, Ramsey is studying the neuroendocrine regulation of food intake. Early in the aging process, obesity is a major health problem, while in very late life, loss of body weight, or wasting, is a major concern. Key neuropeptides in the hypothalamus appear to play a role in regulating food intake. Ramsey is investigating how several peptides—among them leptin, orexin, neuropeptide Y and melanin concentrating hormone—regulate food intake, energy expenditure and body composition in primates. This knowledge could lead to the design of better treatments to either prevent obesity or encourage weight gain.

Jon Ramsey received his bachelor of science degree in biochemistry from Iowa State University. In 1995, he received his Ph.D., in animal sciences from Colorado State University. He completed post-doctoral training at the Wisconsin Regional Primate Research Center in 1998 and is now principal investigator on two NIA grants and one pharmaceutical company grant.

References:
Ramsey, J.J., Harper, M-E., and Weindruch, R. 2000. Restriction of energy intake, energy expenditure, and aging. Free Radical Biology & Medicine. 29:10;946-968.

Ramsey, J.J., Roecker, E.B., Weindruch, R., and Kemnitz, J. 1997. Energy expenditure of adult male rhesus monkeys during the first 30 months of dietary restriction. American Journal of Physiology 272(4, pt 1): E901-E907.

Ramsey, J.J., Kemnitz, J.W., Colman, R.J., Cunningham, D., and Swick, A.G. 1998. Different central and peripheral responses to leptin in rhesus monkeys: Brain transport may be limited.
 

National Laboratory Animal Technician Week recognizes staff improvements to animal care

Animal caretakers and supervisors enjoyed pizza from Drs. Darrel Florence and Joe Kemnitz  to recognize their dedication in providing optimum care for Primate Center colonies. The Feb. 2 lunch celebrated National Laboratory Animal Technician Week, initiated by the American Association for Laboratory Animal Science (AALAS).

Center caretakers have more than 334 years of experience combined in providing excellent animal care. “They can be proud of their many achievements over the past year,” says Darrel Florence, D.V.M., M.Sc., associate director for Animal Services. 

“Animal Services staff meet regularly and are constantly finding new ways to make the environment for the animals better both psychologically and socially,” says Florence. “The best thing we can provide is social housing. The next best thing is challenging foraging opportunies.”  Toward this goal, he and the staff recently worked with Bruce Pape and Duane Zweifel in the Shop Unit to develop a new type of foraging board. “We now have something that can be left on the cage, sanitized with the cage, and is easy to service,” he explained. “Our goal is for every animal to have frequent foraging opportunities.”

Staff development is another area of progress at the center. Most recently, Stacey Maves received her AALAS Laboratory Animal Training (LAT) certification Dec. 5.  “We are all delighted that Stacey took advantage of this great opportunity,” Florence says.