Developmental Effects of Choline: Food for Thought?

Christina L. Williams, PhD

Christina L. Williams, PhDIt is difficult to imagine how the food we eat could alter our brain and lead to changes in behavior. However, since the pioneering work of Curt Richter, we have known that nutrient balance is regulated by both behavioral and physiological processes. More recently, the work of Richard Wurtman has shown how the food we eat modifies our brain, which leads to changes in behavior. One nutrient that has come under considerable scrutiny is choline, best known as a precursor for the biosynthesis of acetylcholine, a neurotransmitter that is involved in learning and memory.

Neurochemical evidence has demonstrated that, under certain conditions, supplementation with dietary choline leads to an increase in transmission of cholinergic neurons and to an improvement in memory function. However, in adult organisms, these memory-enhancing effects of dietary choline are small, transient, and most significant in organisms suffering from choline or acetylcholine deficits. In the adult, there is, as yet, no strong evidence that increases in choline availability can enhance performance above a 'normal' level. Can memory be improved in average individuals by food supplements? If so, when do they need to be taken and under what circumstances are we likely to be vulnerable to deficits in these compounds? These are the questions that my colleague and collaborator, Warren Meck, and I have been investigating over the last 6 to 7 years.

The starting point for our research was the realization that the demand for choline is much greater in the developing organism than in the adult, and that the neonate is likely to be exposed to large changes in the availability of choline. Fetuses and infants require large amounts of choline to make the phospholipids that form the structural backbone of all biological membranes and that are used in transmembrane signaling and methyl-metabolism. Nature has developed several mechanisms to ensure that the developing mammal receives adequate supplies of choline. Choline is actively transported from mother to fetus across the placenta and through the mother's milk. Both the placenta and the mammary gland concentrate choline from the mother's blood serum. Choline concentration is normally much greater in fetal rat blood serum than in maternal blood serum (100 mmol/L vs. 12 mmol/L), and both human and rat milk contain 1.5 mmol/L choline and choline esters. Choline availability to the pregnant and lactating rat determines choline availability to fetal and suckling rats. There is no question that choline intake by the mother determines how much choline the brain of the developing rat can access.

These findings led us to examine the long-term effects of dietary supplementation with choline to rat mothers and developing infants. We have found that supplementation of choline to mothers during specific periods of pre- and postnatal development results in profound improvements in memory function in the pups that last a lifetime. This memory-enhancing effect of choline can be detected in prepubertal rats, is apparent in young adults, and can still be seen in elderly rats at 26 to 28 months of age.

In our studies, choline is administered prenatally to pregnant rats via their drinking water and postnatally via injections. This dose provides, approximately, a five-fold increase in choline intake for the pregnant rat and developing offspring, compared with that provided by standard rat chow. The five-fold range in choline consumption can be easily achieved in human populations.

Diets based on bread, pasta, and potatoes contain less choline than diets that include meat and eggs. For example, 100 g of steak contains the same amount of choline as 1,000 g of potatoes. Thus, the dose of choline administered in our studies is within the range of daily variations in intake that an omnivore like a rat or human might experience. In our studies, no further choline supplementation is given after pups are weaned at 30 days of age.

We have found that choline supplementation during both pre- and postnatal development causes significant and long-lasting facilitation of memory capacity and precision in rats of two strains (Sprague-Dawley and Long-Evans) as assessed using radial-arm maze and water maze tasks. Prenatal choline-treated rats show a greater improvement in visuospatial memory scores, as a function of training, compared with control rats. They are also able to retain a larger number of spatial locations in working memory than are control rats.

A most astonishing finding is that the perinatal choline-treated rats are more accurate on spatial memory tasks even at asymptotic performance levels, suggesting that the treatment may be improving performance beyond the normal range.

We also have demonstrated that choline supplementation during prenatal development has other important effects on visuospatial memory.

Until 20 days of age, control rats are not able to learn the location of a hidden escape platform in a water maze pool by using relational cues. In contrast, prenatal choline-supplemented rats are able to do this task 3 to 4 days earlier. This suggests that the early nutritional supplement may be altering the rate of maturation of memory systems. Perhaps, most surprising, is the discovery that aged rats (25 to 26 months of age), given supplemental choline around the time of birth, show significantly less forgetting of food sites or escape locations when a delay is interposed between training and testing. Like elderly humans, aged control rats have a difficult time holding information in memory during a long delay. These data suggest that early choline supplementation may inoculate against the normal, age-related decline in working memory. In rats, choline supplementation clearly enhances memory function across the entire lifespan.

The next step in our research plan was to determine the mechanism by which perinatal choline exerts these effects. First, we discovered that there are sensitive periods when the brain is particularly responsive to choline supplementation. The first occurs during embryonic days 12 to 17 (pregnancy in rats lasts 22 days), and another occurs during postnatal days 16 to 30. The prenatal sensitive period corresponds to the time when cholinergic cells of the basal forebrain (i.e., a group of cholinergic cells involved in memory processes) are being born. The postnatal period corresponds to the time when cholinergic cells from the basal forebrain undergo synaptic remodeling as they connect to the hippocampus and neocortex. This suggests that our treatments may be targeting a select population of cells. Second, we have begun to collaborate with Dr. Steve Zeisel, Dr. Kryzsztof Blusztajn, Dr. Rebekah Loy, and Dr. Scott Swartzwelder to determine the metabolic fate of our supplemented choline, as well as the long-term neurochemical, neuroanatomical, and neurophysiological consequences of prenatal choline supplementation. Our hope is to discover the biological underpinnings of our memory improvement.

When the fate of the supplemented choline is tracked, Dr. Zeisel and colleagues have discovered that, after 6 days of prenatal choline supplementation to the pregnant rat, fetal brains have a higher concentration of the choline metabolite phosphorylcholine.

Phosphorylcholine is a major storage pool for choline and is the precursor for the synthesis of phosphatidylcholine, which is the major phospholipid in cell membranes. Thus, our data suggest that we may be aiding in cell formation in fetal brains, perhaps by increasing the availability of the substrate for cell membranes. In collaboration with Dr. Loy, we have found that cholinergic cell bodies in the basal forebrain are bigger and rounder in choline-supplemented rats than they are in untreated littermates, suggesting that, perhaps, by providing substrate for cell membranes, we have altered cells within the memory system itself. More important, noncholinergic cells in the basal forebrain are not altered by our choline supplementation. Thus, our effects may be specific to cells that manufacture acetylcholine.

In order to determine if these changes in cell structure translate into changes in the physiology of cells, we have begun to examine a phenomenon called long-term potentiation in the hippocampus of choline-supplemented and control rats. Long-term potentiation is hypothesized to be a possible model for the cellular basis of memory. With assistance from Dr. Swartzwelder's laboratory, we have found that long-term potentiation can be elicited with a considerably lower threshold in hippocampal slices from rats given choline supplementation early in development. It is simply easier to get a potentiated response in these rats; however, when potentiated, there is no difference in the size or duration of the potentiated response.

Together, these data provide strong support for the view that perinatal choline supplementation causes long-lasting changes in the structure and function of the basal forebrain cholinergic system and its projections to the hippocampus and that these changes are likely candidates for the neural substrates underlying the enhanced visuospatial memory.

These studies are the first to demonstrate long-lasting improvements in memory capacity and precision above normal levels of functioning, following a brief period of choline administration. These results reveal that the effects on later behavior appear to be dependent on the timing of choline exposure in early life. Hence, choline appears to be an important, environment-dependent, local organizer of the brain.

As yet, we have no evidence that choline supplementation would improve human memory if given during early development; nevertheless, it is important to consider that human babies certainly are exposed to diets with varying levels of free choline and other choline compounds. Postnatally, human infants may drink breast milk, which is dramatically affected by maternal nutritional status, or a milk substitute. Infant formulas and human milk vary greatly in their total choline content and in their make-up from choline-containing compounds. Some formulas contain only half the total content of choline and choline compounds that human or cows' milk contain. Formulas made from soy protein contain much less free choline and twice as much phosphatidylcholine than does cows' or late lactational human milk. These observations are important because they suggest that the choline content of the human infant diet varies greatly.

Our studies indicate the importance of monitoring the choline content of the mother's diet during prenatal development of the fetus and the choline content of the infant's diet during early postnatal development. Although not certain, our research suggests that choline availability to the fetus and young infant may influence memory development, adult memory capacity, and resistance to age-related memory impairments.

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