According
to a 1994 report of the National Center for Health Statistics,
between 44,000 and 88,000 babies born in the United States each
year may have been exposed prenatally to cocaine during periods
of critical brain development.
The popular antidepressant Prozac is widely prescribed during pregnancy, despite the fact that its effects on the fetus are not well documented. What are the implications for the future generation?
Studying the Effects
of Prenatal Drug Exposure --Through the Minds of Mice
by Mary Beth Zeman
Two William Paterson University biologists - Dr. Martin Hahn, a behavioral geneticist, and Dr. Robert Benno, a neuroscientist - have spent the last decade studying the impact of prenatal cocaine exposure, and, more recently, Prozac exposure, on laboratory mice. Their goal is to establish exactly how those drugs create chemical changes in the brain and what the impact is on behavior.
"Drug use in pregnant females carries special risks for a developing fetus," says Hahn, noting that significant physical and behavioral consequences, including neurologic problems, growth retardation, sleep disorders, and aggression have been observed in human newborns and infants exposed to cocaine. "What is not well understood are the precise neurobiological changes that occur in the brain, and how those effects are related to changes in behavior."
The pair are particularly interested in the long-term outcomes of drug use - whether illicit or licit - during pregnancy. "A number of studies, including our own, indicate that prenatal exposure may produce very subtle differences, perhaps associated with higher-level functioning skills such as problem solving," Benno says. "Under a stressful social situation, for example, more aggressive behavior might be exhibited."
The brain is an incredibly complicated organ. While the human brain is more complex and highly developed than that of any other animal, the brains of all mammals are remarkably similar.
Averaging three pounds
in a human adult, the brain contains between 50 billion and 200
billion information-processing cells, called neurons. They have
two unique appendages: axons, from which nerve signals are transmitted,
and dendrites, which receive messages from other neurons.
Aiding the transmission of messages across the synapse, or gap
between two neurons, are a variety of chemicals called neurotransmitters,
which are stored in small vesicles in the ends of the axons.
When a nerve impulse is sent, the axon releases its neurotransmitters
into the synapse, where they bind to receptors typically located
on the dendrites.
Cocaine has been shown to operate on three specific neurotransmitters: dopamine, which among other things controls pleasure and behavioral flexibility; serotonin, which is implicated in appetite and behavioral satiety; and norepinephrine, which impacts arousal. "Specifically, cocaine works by blocking the reuptake, or reabsorption, of these neurotransmitters," Benno explains. "As a result, there is an increase in the functional amounts of these chemicals in the brain - and, essentially, a dysfunction in the behaviors these neurotransmitters regulate." For example, arousal refers to one's state of wakefulness. "An arousal dysfunction could range from someone who is manic to someone who is incapable of staying awake," he notes.
Since it would be difficult to run a controlled study of how these drugs impact on humans, scientists such as Hahn and Benno turn to animal models in which the effects on offspring can be explored. With funding through the College of Science and Health's Center for Research, the pair have used mice to perform a number of experiments in an attempt to dissect the impact of drugs on the brain and behavior.
The mouse lab is located on the lowest level of Science Hall, behind a locked door in a quiet hallway. Norman Schanz, the Biology Department's animal caretaker, keeps a watchful eye on his tiny charges, which number from 200 to 3,000, depending on the needs of a particular experiment. The mice live in clear plastic cages equipped with food and water which is always available.
The mice are bred on campus, allowing the scientists to apply genetic control to their experiments. "By using a particular inbred strain, we can produce mice that have virtually identical genetic profiles," says Hahn. "This allows us to look not only for behavioral or chemical changes caused by the drugs, but also to determine whether the impact of the drug varies by genotype."
Mice are typically
born after 21 days. At the age of 45 days, they are considered
adults. This time-compressed span of development is one of the
main reasons the researchers believe mice are excellent subjects
for this project. "Within a relatively short period of time,
we can observe the mouse's behavior from birth to adulthood.
A similar study of human behavior would require more than two
decades," Hahn explains.
(Photo: Drs. Hahn and Benno with a few of their charges.)
One recent experiment looked at the level of aggression in adult male mice with three distinct genetic profiles. Female mice were mated with one of the three genetically distinct males. Once the mating was successful, the pregnant females were isolated and randomly divided into two groups. During days seven to 17 of gestation, each group received a daily injection - one was given a carefully calculated dosage of cocaine; the second received saline. "A side effect of cocaine can be weight loss," Benno explains. "The process of selecting a dosage that will not cause the fetus to be malnourished is critical, thus allowing us to study the direct impact of the drug."
Male pups born after prenatal exposure were tested at about 80 days of age. Two males were placed in circular nest boxes attached by a tube with a gate at each end; the mice could smell each other but could not engage in physical contact. After three hours, the gates were removed and the mice were free to interact. Aggressive behaviors such as upright boxing, chasing, fighting and squeaking were logged in real time.
Hahn and Benno differ in their interpretations of the data. "In my view," Benno explains, "prenatal cocaine exposure limited the animal's ability to respond appropriately to its environment. The mouse fought the same way, whether he faced a more passive opponent or a more aggressive one."
In Hahn's view, the drug-exposed mouse was more predatory. "If the cocaine-exposed animal was presented with a non-aggressive mouse, he attacked; yet if he faced a more aggressive mouse, he backed off," he says. However, regardless of their views of the data, both scientists agree cocaine exposure changed behavior in this social situation.
In another experiment focusing on social behavior, the researchers examined the ultrasonic sounds made by the newborn pup. "In mice, auditory signals produced by pups play a critical role in the regulation of the mother-infant relationship," Hahn explains. "Certain calls reliably elicit the mothers to retrieve their young for care and feeding."
Again, half the pregnant females were injected with cocaine, the other half with saline. On days two through four, the pups were briefly isolated from their mothers and their vocalizations were recorded. In two of the three genotypes studied, the cocaine-exposed pups called less frequently during their early days.
"This is significant,"
Hahn points out. "Newborn mice are unable to walk or see.
By vocalizing, they motivate the mom to search for them and find
them. If the mom does not retrieve them, they face death from
hypothermia." Their results mimic those of a recent study
of human infants, which found that cocaine-exposed babies cried
less and for shorter duration.
"Both of our studies point to a small, but reliable, impact
of prenatal cocaine exposure," Benno says. "The effects
are subtle, but they are there, and thus provide us with the
motivation to continue our research and seek an even deeper understanding
of these issues."
Now,
the scientists plan to add Prozac to their experimental mix.
The reason is twofold. "Prozac is known to affect only one
neurotransmitter, serotonin," Benno notes. "By comparing
how mice exposed to cocaine and Prozac react, we can gain a better
understanding of cocaine's impact on the brain's neurotransmitter
systems and how specific effects on individual transmitter systems
might be related to behavior."
Secondly, Benno believes it is important to study Prozac in its own right. "Prozac is a class C drug in terms of pregnancy, which means that doctors can prescribe it if the benefit to the mother outweighs the risk to the fetus," he says. "To me, it is societally important to try and understand what impact such a drug might have. The changes might be negligible in infants - but the real question is what impact such prenatal exposure will have on someone 20 or 30 years from now."
Last summer, the pair worked to determine the correct dosage for Prozac, one that will not affect other factors such as weight gain in the pregnant mice. This year, they will embark on a comparison of cocaine- and Prozac-exposed mice. "We plan to study brain and body growth, sensory-motor development, and the ultrasonic calls of female mice," Benno explains.
Both scientists say they have truly enjoyed working together as collaborators. "This project came about initially through a brainstorming session," Hahn reveals. "We each enjoy having someone to bounce ideas off of. We're also both curious, which is a strong motivator."
Each brings a unique perspective - in terms of personality style, teaching approach, and expertise - to the research. Hahn, who describes himself as "very disciplined and more linear," has studied animal behavior for nearly 30 years. A graduate of Ohio State University with a bachelor's degree in psychology, he earned his master's degree in psychology and his doctorate in experimental psychology from Miami University of Ohio. Prior to joining the William Paterson faculty in 1973, he spent three years as a National Science Foundation research associate studying behavioral genetics with John Fuller, who is considered one of the founders of the field. He and his wife, Lynn, have two grown children, Eric, 28, and Sarah, 25.
Benno has long been interested in the regulation of neurotransmitters and enzymes in the brain. A graduate of Tulane University with bachelor's and master's degrees in chemistry, he earned a doctoral degree in anatomy and neuroscience from the University of Iowa in 1978. He joined William Paterson in 1982 after four years as a postdoctoral fellow and instructor in the Laboratory of Neurobiology at Cornell University Medical College in New York City, where his research focused on the development of a computer-assisted quantitative immunocytochemical method to study brain chemistry. "I don't rein in my curiosity quite as well as Marty does," says Benno. "If the research results in something interesting, I'm more likely to want to travel down a new path, whereas Marty will want to stay on track." He and his wife, Karen Donelson, a physical therapist, have a six-year-old daughter, Jia Mae, "who loves to come to the lab and see daddy's baby mice."
In addition to their
research, which has generated numerous journal articles and book
chapters, the pair have served as co-editors, with J. K. Hewitt
and H.D. Henderson, of Developmental Behavior Genetics: Neural,
Biometrical, and Evolutionary Approaches (Oxford University Press
1990). Benno also serves as coordinator of the University's Honors
Program in Biopsychology, which Hahn was instrumental in creating
in 1976.
"There is a deal of creative tension involved in our collaboration,"
Hahn says. "But I believe we work on each other in a good
way. We represent two different approaches, and over the years
have been able to appreciate the other's perspective."
One thing both agree on is the importance of having students work with them in the lab. Throughout the years, they have supervised dozens of students, particularly undergraduates, who have participated in data collection on a variety of projects (see sidebar).
"We look forward
to the summer when we can spend two months solely focused on
the research and have the opportunity to work with students,"
says Hahn. "Because this project involves data collection
seven days a week, the students play an integral role. We couldn't
operate without them." During the summer of 1998, they supervised
seven students, with support from the Center for Research, as
well as two minority students through a mentorship program, Pathways
to Academic Success in the Sciences, administered by the University's
Office of Minority Education.
"Our greatest reward is to see our students be successful,"
Benno adds. "The Biology Department as a whole is committed
to the value of undergraduate research and we enjoy giving that
opportunity to students through our projects."
Through funding from the University's Center for Research, Hahn and Benno have been able to explore an area of research that captivates them both. "We are incredibly grateful to the institution for its continued support of our efforts," Hahn says. Grants from the Center are awarded via a competitive process with reviewers from outside the University.
Motivated by their interests in the brain and behavior, Hahn and Benno are committed to continuing their research. "We're becoming a more medicated society," Benno says. "We know that drugs, whether illicit or licit, alter the neurochemical composition of the brain. By gaining a better understanding of these changes and their impact on behavior, we can then develop strategies for assisting drug-exposed individuals in solving problems they will face down the road."WP
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Animal Research Subjected To Strict Guidelines All William Paterson researchers seeking to use animals in experiments must follow strict guidelines spelled out by the National Research Council that are designed to promote humane care of animals used for biomedical and behavioral research, teaching, and testing. The guidelines are
enforced by University's Committee on Laboratory Animal Welfare,
which by law must include a doctor of veterinary medicine, at
least one practicing scientist experienced in research with animals,
and at least one public member to represent general community
interests. The committee, appointed by the provost, meets twice
each academic year to review proposals, discuss relevant issues,
The facilities are also subject to random inspection by the U.S. Department of Agriculture's Animal and Plant Health Inspection Service. "We have always complied with U.S.D.A. and National Institutes of Health regulations and have been cited for the high quality of care we give our animals," says Marty Hahn. |
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Research Participation Spurs Student Project
For psychology major Christopher Hess, working in the mouse laboratory with biology professors Marty Hahn and Robert Benno has been "an incredible opportunity," one that has led to an independent study project and cemented his interest in neuroscience. Hess joined their research team during the spring of 1998. "I was involved in data collection - measuring and weighing the animals, weighing the food they consumed, and calculating the injection dosage for Prozac, which is a new element to the study," he explains. He noticed some results which were contrary to current scientific thought. "Prozac is known to decrease food consumption," he says. "Instead, I found that the mice were increasing their food intake." He approached Hahn and Benno about his findings, and was encouraged to begin an independent study project. Throughout the summer and into the fall, he examined other variables such as method of injection and genotype; neither had any impact on the results. "Each finding leads me to another hypothesis," he says. "This process has given me a real understanding of how research works and what the implications are." Admittedly a less than motivated student in high school, Hess enlisted in the Marine Corps after he graduated from Indian Hills Regional High School in Oakland in 1992. During his four-year tour of duty in the security forces and infantry divisions, he was stationed in California, Alaska, Hawaii, and Japan. He picked William Paterson for its proximity to home, and has found his time on campus invaluable. "Most undergraduates would not have the chance to conduct a research project like mine," he says of his experience. A junior who is also enrolled in the biopsychology honors program, he plans to pursue a doctoral degree in psychology or neuroscience. |
n
many colleges and universities, the opportunity to participate
in significant research projects is limited to the graduate level.
At William Paterson, and particularly in the Biology Department,
undergraduates are encouraged to join faculty in their research
initiatives as part of a commitment to developing student involvement
through experiential learning.