Neurobehavioral assessment during the early postnatal period

Neurotoxicologyand Teratology,Vol. 12, pp. 489-495. ©Pergamon Press plc, 1990. Printed in the U.S.A.

0892-0362/90 $3.00 + .00

Neurobehavioral Assessment During the Early Postnatal Period LINDA PATIA SPEAR 1

Department of Psychology and Center for Developmental Psychobiology SUNY-Binghamton, Binghamton, NY 13901

SPEAR, L. P. Neurobehavioral assessment during the early postnatal period. NEUROTOXICOL TERATOL 12(5) 489-495, 1990. --Few laboratories investigating the neurobehavioral consequences of developmental toxicants assess offspring early in ontogeny other than examining physical maturation, reflex development and perhaps locomotor activity, measures which tap only a limited portion of the neurobehavioral capacities of young organisms. The importance of including a wider range of neurobehavioral assessments during the early postnatal period in developmental toxicology test batteries is discussed. Special considerations for the design of testing early in life are enumerated, and examples are given of suckling, cognitive and psychopharmacological tests that have been shown to be sensitive indicators early in life of the effects of gestational drug exposure. Developmental toxicology Psychopharmacology

Neurobehavioral teratology

Ontogeny

Neonates

Suckling

Cognitive

from clinical studies of young humans, commonalities should be greater when laboratory animal populations are similarly examined early in life [see (40) for further discussion]. In addition to specific effects on nervous system development, exposure to a developmental toxicant may have a number of nonspecific effects (such as alterations in maternal hormone release, hypo- or hyperthermia, hypoxia, alterations in nutritional status, etc.) that may also have an impact on nervous system development. Primary (neural) effects of a developmental neurotoxicant may subsequently lead to various secondary disruptions in the development of other neural systems. Thus, exposure to a developmental toxicant may not result merely in a simple set of neural alterations that are maintained throughout life, but rather may alter the long-term prospect for future development, leading to a concatenation of alterations in neurobehavioral functioning that are gradually elaborated as the animal matures. The earlier that testing occurs following the time of chemical insult, the more likely that primary alterations induced by the insult will be evident. When attempting to develop principles regarding or governing the types of neural alterations that may be induced by specific classes of developmental neurotoxicants/behavioral teratogens, it may be important to focus initially on primary alterations rather than the full complement of consequent reactions to such an agent. Assessing the consequences of a potential neurobehavioral teratogen only in adult offspring may not reveal transient deficits, even though such transient alterations may have profound longterm consequences for the organism. In some cases, substantial recovery of function may occur with time [see (22) for further discussion], although it is also possible that in other circumstances

ALTHOUGH the importance of age at testing for assessing the effects of developmental toxicants has been periodically recognized (22, 24, 32), the vast majority of neurobehavioral assessments are still conducted on adult animals. Yet, there are a number of advantages to including assessments of neurobehavioral function early in life. This paper will first discuss the advantages of such early assessments and then will address the issue of how tests can be effectively designed for examination of neurobehavioral function early in life. The focus of this presentation will be on the early testing of altricial rodents, although the principles outlined may also be applied to the young of other species. ADVANTAGESOF EARLYASSESSMENT There are numerous advantages of early assessment. On a pragmatic level, such testing can be cost effective. Tests can be chosen that are easily and rapidly conducted. Most importantly, maintenance costs are substantially less than when testing exposed offspring as adults. Even when conducting work that mandates adult testing (such as "segment 1" testing), all animals in a litter may not be ordinarily maintained until adulthood for cognitive and reproductive testing. By conducting neurobehavioral assessments early in life on offspring that would normally be discarded at weaning, additional data can be obtained without an impact on maintenance costs for the overall project. Early testing is not only cost effective, but can also be important scientifically. Most clinical studies of early toxicant exposure are conducted on human infants and young children. Yet typical studies of developmental toxicants in laboratory animals focus on neurobehavioral assessments in adulthood. When comparing data obtained from laboratory animal studies with those

~Requests for reprints should be addressed to Dr. Linda Spear, Department of Psychology, SUNY-Binghamton, Binghamton, NY 13901.

489

490

SPEAR

TABLE 1 APPROXIMATE AGES DURING WHICH VARIOUS SENSORY AND MOTOR CAPABILITIES ARE EVIDENT IN RAT PUPS

Postnatal Day:

0

Sensory capabilities

7

14

21

olfactien¢ scmatosensory (tactile)~ thermal r vestibular r taste

)

auditory

( Motor capabilities

<

crawl a

(

piv°ting c

<

probing

)

walking

)

<

d

>

visit

)

b

>

==~g

>

> wall climbing

) <

supported rear (

face washin~

)

(

> rear

>

emergence of other aspects of 9"roaning

)

~xplor.e solid food

)

sucklin@ e

)

~Crawl= forward movements with stomach on ground. bWalking= forward movements with stomach off ground. cPivoting = turn to L or R propelled by forelimbs; hindlimbs stationary. dProbing = pushing with snout against solid object. ~l"hroughout the suckling period, pups can independently ingest liquids when puddled on the floor or when infused into the oral cavity. particular functional deficits may not emerge or become evident until later in life [see (16) for examples]. For instance, a developmental toxicant could produce pronounced cognitive deficits early in life, with substantial recovery occurring thereafter. Yet, transient disruptions in cognitive function early in life, when much critical learning normally occurs, may have a long-term impact on the individual, even if normal cognitive function were to emerge later in life. CONSIDERATIONS FOR THE DESIGN OF EARLY NEUROBEHAVIORAL TESTING

Typical test batteries used for initial screening of potential behavioral teratogens include a number of measures designed to assess particular categories of functional effects. For instance, the following categories of functional effects were recommended for inclusion in initial screening tests by a 1985 work group (8): physical growth and maturation, reflexes, motor development/activity, sensory/attentional function, affect, cognition (learning/memory/performance), reproductive behavior. Only the first three of these categories are frequently assessed in young animals. Assessments of physical growth and maturation are typically indexed by body weight and the time of reaching various landmarks of physical maturation (eye opening, upper and lower incisor eruption, etc.). Reflexes are assessed by their developmental age of appearance, and motor development/activity assessments sometimes include testing early in life. These tests, although important, assess only a limited portion of the neurobehavioral repertoire of young organisms. In principle, all other categories of functional effects except reproductive behavior can

also be examined early in life, although in practice only a few laboratories have begun to do so. Researchers unfamiliar with young organisms sometimes consider neonates to be behaviorally impoverished organisms with only rudimentary sensory capacities. This is simply not the case. Altricial newborns such as rat pups are capable of exhibiting a wide repertoire of motor movements under environmentally appropriate circumstances, and of demonstrating exquisite sensitivity to certain modalities of sensory stimuli. For instance, Table 1 lists the approximate ages at which certain sensory capabilities and motor responses are present in rat pups. With appropriate consideration of the special capabilities and limitations of these young animals, many of the same types of behavioral tests used to assess various categories of functional effects in adult animals can be modified for use early in ontogeny. For example, although it is not possible to use auditory or visual stimuli until the late preweaning period, rat pups are very sensitive to olfactory, taste, tactile and thermal cues from birth or before [see (16) for further discussion and references]. In terms of thermoregulation, rat pups are poikilothermic and only gradually develop thermoregulatory capacities during the late preweaning period; during this time, the behavior of rat pups is often markedly influenced by body temperature [e.g., (18)]. Test protocols can easily be modified, however, to allow for testing pups in temperature-controlled situations and for the systematic monitoring of pup body temperature. Behavioral assessments can often be effectively modified to consider the special response capabilities of young rat pups. For instance, neonatal rat pups have low baseline levels of motor movements which could be problematic in certain types of testing situations. As one example, this low baseline response rate presents difficulties when attempting to assess psychopharmacological responsiveness to a drug expected to depress motor

EARLY NEUROBEHAVIORAL ASSESSMENT

movements. There are ways to circumvent this problem. For instance, if rat pups are deprived of the dam (and hence food) for several hours, they exhibit a marked increase in behavioral activation when exposed to milk (either via intraoral infusions or by puddling milk on the floor) in a warm apparatus (9,10). By testing pups in both the absence (low activity baseline) and presence (high activity baseline) of milk, it is possible to detect both increases and decreases in behavioral responses to pharmacological challenges in young rat pups [e.g., see (33)]. Some types of conditioning situations require very minimal modification for testing early in life. For instance, classical conditioning procedures have proved to be particularly useful for cognitive testing in infants, given that preference tests mandating only small movements to one side or other of an apparatus can be used as a measure of conditioning [e.g., see (36)]. Other types of cognitive tests used in adult animals can also frequently be modified to consider the special response capabilities of young rat pups. For instance, an operant bar press response used in adults can be modified to an operant head probe/head lift response for neonates [e.g., (17)]. Indeed, under appropriate circumstances, neonatal rat pups are capable of demonstrating conditioning of an instrumental discrimination task. Johanson and Hall (17) used the tendency for rat pups to become activated and probe upward in the presence of milk to condition an appetitive operant discrimination response in newborn rat pups. One-day-old rat pups learned to probe into a terry-cloth-covered paddle when paddle deflection was rewarded with oral milk infusions. In a two-paddle situation, if different odors were placed on the two paddles, the neonates learned a lever-press discrimination, i.e., to probe selectively the paddle that was paired with milk. EXAMPLES OF EARLY NEUROBEHAVIORAL TESTS

As discussed above, testing of infant animals can be designed to examine the same types of functional categories that have been typically examined in adult animals. Some examples of the types of testing that can be conducted in young animals and their usefulness in detecting behavioral alterations following exposure to developmental toxicants follow.

Age-Specific Behaviors of Infancy: Suckling Behavior There are a number of age-specific behaviors of infancy that appear to be strongly linked to the functioning of particular neurotransmitter systems. Among these behaviors is suckling, an essential age-specific behavior that appears to be dependent on serotonergic (and cholinergic) activity particularly during the first postnatal week [e.g., (23,30)]. Suckling can be easily assessed by placing pups in the presence of an anesthetized dam and measuring suckling attachment latencies, length of time attached, and even suckling pressure or number of sucks on the nipple (through the use of pressure transducers and electromyographic recordings, respectively). Suckling test procedures appear to be sensitive indicators of gestational drug exposure. For instance, Riley and associates (6) observed suckling deficits in 6-7-day-old offspring exposed gestationally to ethanol, both in terms of the amount of time spent attached and the degree of negative pressure exerted on the nipple; these effects were only transitory in that no such deficits were seen at 9-10 days of age. These data are reminiscent of those observed in human infants exposed gestationally to ethanol who have also been reported to exhibit transient deficits in suckling during the early postnatal period [e.g., (41)]. It is interesting that among the neurochemical alterations reported in rodent offspring exposed gestationally to ethanol are permanent attenuations in, or delays in the development of, cholinergic and serotonergic activity (4, 5, 13). Hence, these behavioral alterations in suckling, which so far

491 have only been revealed by testing during the early postnatal period, are consistent with clinical reports as well as with predictions based on the neural substrates affected by this developmental toxicant.

Cognitive Assessments During the Early Postnatal Period Classical conditioning tasks have been shown to be sensitive to the effects of gestational drug exposure. For instance, we have observed that 7-8-day-old infant rats exposed gestationally to cocaine do not exhibit significant conditioning in an appetitive classical conditioning task which is reliably learned and retained for 24 hr by control offspring (34). In this experiment, deprived pups exposed gestationally to cocaine, or pups derived from dams pair fed to the cocaine-treated dams (PF) as well as untreated dams (LC), were given three massed training trials. Each trial consisted of a 3-min exposure to a chamber containing a conditioning stimulus (odor) that was not paired with any explicit reinforcement ( C S - exposure), 3-min exposure to a chamber containing a conditioning stimulus (odor) which was paired with reinforcement via intraoral infusions of milk (the unconditioned stimulus, US) every 30 sec (CS+ exposure), followed by 3 min in a holding cage. Other pups from each prenatal treatment group were placed into an unpaired condition; these animals received milk exposure prior to the presentation of the odors. Testing occurred immediately and 24 hr after training, with each 3-min test consisting of a spatial preference test between the CS + and CS - odors. For the test, the two odors were placed on cotton located underneath the apparatus floor at opposite ends of the test apparatus; each pup was placed on the midline of the apparatus and the amount of time spent on each side of the chamber recorded. Animals of this age are easily able to move the small distances required to exhibit a preference in this test situation. As can be seen in Fig. 1, paired LC and PF offspring spent significantly more time on the CS + side of the apparatus than their unpaired counterparts, thus exhibiting significant conditioning in this task. No sign of conditioning was seen in the offspring exposed gestationally to cocaine. We have also observed similar conditioning deficits in offspring exposed to cocaine during gestation when they were trained at 17-18 days of age in an aversive classical conditioning task where CS+ odor was paired with brief footshock exposures (35). Deficits in classical conditioning tasks are not always seen, however, in cocaine-exposed offspring tested early in life. Although we have yet to examine this systematically, a critical factor appears to be the degree of training on the task, with cocaineexposed animals being less likely to exhibit conditioning deficits when trained using procedures that result in strong conditioning. For instance, no conditioning deficit was seen in 7-day-old cocaine-exposed offspring tested on an aversive classical conditioning task where the training consisted of 3 CS+/US trials each of which lasted 3 min, with footshock being delivered for 3 sec twice during each CS+ odor exposure (35). Yet, as can be seen in Fig. 2, 8-day-old offspring exposed gestationally to cocaine did exhibit conditioning deficits in an aversive classical conditioning task where pups were given only two CS+/US trials, each of which lasted only 20 sec during which two 3-sec footshocks were delivered (14). In both of these studies, preference tests as outlined above were used as an index of conditioning. Classical conditioning assessments conducted early in life also reveal cognitive deficits in offspring exposed gestationally to ethanol. For instance, 3-day-old ethanol-exposed offspring did not exhibit conditioning in an appetitive classical conditioning task (where the CS + odor was paired with intraoral milk infusions as the US), whereas offspring of both pair-fed and ad lib dams did exhibit significant conditioning in this task as indexed by an increased preference for the CS+ odor (measured using preference

492

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ODOR PREFERENCE(IttlEDIATE)

PAIRED

[]

UNPAIRED

140

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120

25 15

0 UJ

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co 0 Z 0 UJ

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80 60

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50 ¸

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FIG. 1. Mean (-S.E.M.) odor preference difference scores [(CS+)(CS-)] for paired and unpaired offspring of cocaine-exposed dams (C) (black bars), pair-fed dams (PF) (hatched bars) and nontreated lab chow control dams (NT) (white bars), immediately and 24 hr following odor/milk conditioning conducted on postnatal day 7. [Reprinted from (34) with permission.]

test procedures similar to those outlined above) in CS + / U S paired pups relative to their unpaired counterparts (1). In this experimental series, performance in an aversive classical conditioning task was also assessed in 10-day-old pups, using an odor as the C S + and illness produced by lithium chloride as the US. Offspring of pair-fed and ad lib control dams that received a paired presentation of the odor and lithium chloride exhibited an aversion for the CS + odor relative to their unpaired counterparts as indexed using the preference test procedure. No sign of conditioning was observed in ethanol-exposed offspring. These investigators also examined aversive classical conditioning in adult offspring exposed gestationally to ethanol and observed no alterations in preference test performance relative to control animals (1). Thus, in ethanol-

FIG. 2. Mean time spent on the CS+ side in sec (-S.E.M.) for paired (white bars) and unpaired (hatched bars) offspring of cocaine-exposed dams (C40) and lab chow control dams (LC) during a 3-min preference test conducted beginning 3 min following odor/footshock conditioning on postnatal day 8.

exposed offspring, cognitive deficits may be more readily observed in testing conducted early in life than in adulthood. Given that cognitive deficits have also been reported in human children exposed gestationally to ethanol (42,43), these data point out the important clinical relevance of testing early in life. Careful selection of the cognitive tests chosen for early testing can be used to address specific questions regarding the nature of cognitive deficits seen in offspring exposed to chemical insults early in life. As discussed above, we have observed deficits in cocaine-exposed offspring in a number of appetitive and aversive classical conditioning tasks in testing conducted during the neonatal to weanling period [see (34,35)]. However, it is not possible to determine from these data whether the cognitive deficits are due to a delay in cognitive development, or to fundamental cognitive deficits per se. Indeed, there are ontogenetic improvements in conditioning with most learning tasks, making it difficult to determine whether impaired performance is due merely to a delay in cognitive development or to specific deficits. There are some types of conditioning situations, however, in which young animals learn more rapidly than older animals. These situations appear to be sensitive to the predisposition of infants to associate different sensory stimuli that are relatively neutral physiologically or to associate a sensory stimulus with particular physiological consequences in a manner that is different from that of the adult [e.g., (37-39)]. We have used one of these tasks, sensory preconditioning, to examine cognitive performance in cocaine-exposed offspring. In this particular version of this task, simultaneous preexposure to two relatively neutral odors was followed by the pairing of one of these odors with footshock. The test for sensory preconditioning consisted of determining whether the animal has developed an aversion to the other preexposed stimulus that was not paired with footshock. Rat pups from 8 to 17 days of age readily exhibit sensory preconditioning in this task, conditioning that is not

EARLY NEUROBEHAVIORAL ASSESSMENT

evident with the same training parameters in weanling (21day-old) animals (14,37). During the preweaning period, cocaineexposed offspring might be expected to perform more poorly on this task than control animals regardless of whether the cognitive deficits in these animals are related to a maturational delay or a fundamental alteration in cognitive performance. However, at weaning, when the ability of normal pups to exhibit this conditioning is dissipating, cocaine-exposed pups would paradoxically be expected to perform better on this task than controls if their deficit is due to a cognitive delay. The sensory preconditioning procedure consisted of three phases. In Phase I (preexposure), each pup was exposed to the CS 1 (banana) and CS2 (lemon) odors simultaneously for 3 min. Forty min later Phase II (conditioning) was conducted. In Phase II each pup was exposed for 20 sec to a C S - odor (almond) followed immediately by placement in a chamber containing the CS2 odor (lemon)• Two brief (3-sec 0.5-mA) footshocks were administered during the 20 sec CS2 exposure. Following a 1-min intertrial interval, pups were given another conditioning trial. Three rain following the second conditioning trial, Phase III (test) was conducted. The test for sensory preconditioning consisted of a 3-min preference test between the CS 1 odor (banana) and a novel odor (orange). To the extent that pups learned an association between banana and lemon in Phase I, when lemon is paired with footshock in Phase II, they should demonstrate an aversion to banana when tested in Phase III. Two types of conditioning controls were used: one which received unpaired exposure to the two odors (CS 1 and CS2) in Phase I, and the other which received unpaired exposure to the CS2 odor and the US (footshock) in Phase II. As can be seen in Fig. 3, paired lab chow control offspring exhibited significant sensory preconditioning (i.e., an aversion to the CS 1 odor relative to unpaired animals) when tested at 8 and 12 clays of age but not at 21 days of age, thus demonstrating the normal ontogenetic decline in performance on this task. Cocaine-exposed offspring did not exhibit significant sensory preconditioning at 8, 12 or 21 days of age. The results of this study (14) are consistent with an interpretation that gestational cocaine exposure causes a fundamental cognitive deficit in these animals, rather than a delay in cognitive development. This interpretation needs to be verified, however, using other cognitive tasks before drawing strong conclusions.

493

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8-DAY-OLD PUPS 120 100 80

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12-DAY-OLD PUPS 140 120 -

Z z m z

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L) uJ <

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100 -

LU

li

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LC

C40

21-DAY-OLD PUPS 140 120

z

< z

100

~

< m z LU I

Virtually all psychoactive drugs elicit behavioral responses in young animals. For instance, Table 2 presents several examples from the work of our laboratory examining the behavioral response to drugs affecting particular neurotransmitter systems. As can be seen in this table, drugs affecting a wide variety of neurotransmitter systems elicit reliable behavioral responses in young animals. In all these cases, behavioral responses were observed from the earliest age tested, although the nature of the behavioral response elicited may not be adult-typical in either qualitative or quantitative terms [e.g., (26-28)]. This is not surprising in that neurotransmitter systems undergo a substantial amount of development during the early postnatal period [see (19) for review]. Even in their immature state, however, these nascent neurotransmitter systems appear to be sensitive to pharmacological manipulations as indexed by reliable behavioral alterations. This psychopharmacological responsivity provides a means to assess functional activity of particular neurotransmitter systems early in life. Examining behavioral responses to acute drug challenges has been shown to be a valuable tool in developmental toxicology. For instance, drug challenges have been used to "unmask" deficits in adult offspring exposed to chemical insults early in life, deficits

UNPAIRED

140

o

Psychopharmacological Challenges Early in Life

[]

z LU

60 40

'° I 2O 0

•

LC

C40 TREATMENT

FIG. 3. Mean time spent on the banana side of the preference testing apparatus in sec ( ---S.E.M.) for paired (white bars) and unpaired (hatched bars) offspring of cocaine-exposed dams (C40) and lab chow control dams (LC). Sensory preconditioning is evidenced by a significant decrease in preference for the banana side of the apparatus in paired animals when compared with their unpaired counterparts. There were no differences in odor preference between animals given unpaired exposures in Phase I versus Phase II, and hence the data have been collapsed across these two unpaired groups in this figure. that may not be evident under baseline testing conditions [e.g., (11,15)]. Moreover, psychopharmacological testing using drugs with specific effects on given neurotransmitter systems can be used to provide initial indications of potential functional alterations in specific neurotransmitter systems, findings which can then be corroborated neurochemically [see (11) for further discus-

494

SPEAR

TABLE 2 PSYCHOPHARMACOLOGICALRESPONSESTO DRUGSAFFECTINGVARIOUS NEUROTRANSMITFER(NT) SYSTEMSEARLYIN ONTOGE,NY

NT System Affected Catecholamine

Serotonin

Acetylcholine GABA

Opiate

Glutamate

Drug

Age When Response Observed*

Behavioral Resp.

Reference

Apomorphine

P7

Locomotor act., wall climbing

(26)

Clonidine

P7

Locomotor act., wall climbing

(28)

Cocaine

P7

Locomotor act.

(28)

Quipazine

P3

Mouthing, analgesia

(29,31)

Methysergide

P3

Suckling, mouthing, hyperatgesia

(7, 30, 31)

Scopolamine

P7

Latency to choicediscrim, task

(27)

Muscimol

P3-4

Locomotor activity

(33)

Picrotoxin

P3-4

Locomotor activity

(33)

Morphine

P3-4

Analgesia

(31)

Naloxone

P3-4

Hyperalgesia

(31)

MK801

P3-4

Locomotor activity

(21)

*In all cases, earliest age tested.

sion]. Psychopharmacological testing is readily adaptable to testing early in life, and may produce valuable data regarding potential neural alterations in exposed offspring. For instance, we have recently examined the psychopharmacological profile of the D2 agonist quinpirole in cocaine-exposed offspring at postnatal day 21 (20). In this study, offspring were examined for 5 min at 30 and 60 min following injection of doses of 0, 0.04, 0.08, 0.5 or 1.0 mg/kg quinpirole; data were collected using a time-sampling procedure in which all behaviors of the pups were observed and recorded for 5 sec every 20 sec during each 5-min test. Offspring exposed gestationally to cocaine were observed to exhibit a shift to the left in the quinpirole dose-response curve for a number of quinpirole-induced behaviors, indicating an increased sensitivity to quinpirole. To the extent that psychopharmacological data are an accurate reflection of alterations in neural substrates, these data suggest that there might be an increase in D2 binding in these offspring. Indeed, we subsequently observed that D2 binding was increased in striatal homogenates of weanling offspring exposed gestationally to cocaine (25). A number of laboratories are beginning to utilize psychopharmacological challenges early in life in developmental toxicant assessment [e.g., (2-5, 12, 20, 35)].

SUMMARYAND CONCLUSIONS It is hoped that these few examples have served to illustrate the utility of testing early in life using measures other than the typical assessments of reflex development and physical maturation. With few modifications that take into consideration the capacities and limitations of young organisms, testing can be modified for infant animals to examine the same types of functional categories as have been typically examined in adulthood. Such testing can be cost effective and, in some instances, may reveal functional deficits that are difficult or impossible to discern in adult test batteries. However, it should be realized that early neurobehavioral assessment alone is not sufficient. Testing throughout the life span is ultimately necessary to discern the effects of any potential developmental toxicant on neurobehavioral function. Thus, early testing should be considered to be an important addition to, not a substitute for, traditional behavioral test batteries in developmental toxicology. ACKNOWLEDGEMENTS This research was supported in part by National Institute on Drug Abuse grants R01 DA04478 and K02 DA00140.

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EARLY N E U R O B E H A V I O R A L ASSESSMENT

macology (Berlin) 96:161-168; 1988. 8. Geyer, M. A.; Reiter, L. W. Strategies for the selection of test methods. Neurobehav. Toxicol. Teratol. 7:661-662; 1985. 9. Hall, W. G. The ontogeny of feeding in rats: I. Ingestive and behavioral responses to oral infusions. J. Comp. Physiol. Psychol. 93:977-1000; 1979. 10. Hail, W. G.; Bryan, T. E. The ontogeny of feeding in rats: II. Independent ingestive behavior. J. Comp. Physiol. Psychol. 94: 746-756; 1980. 11. Hannigan, J. H.; Blanchard, B. A. Commentary: Psychopharmacological assessment in neurobehavioral teratology. Neurotoxicol. Teratol. 10:143-145; 1988. 12. Hannigan, J. H.; Blanchard, B. A.; Homer, M. P.; Riley, E. P.; Pilati, M. L. Apomorphine-induced motor behavior in rats exposed prenatally to alcohol. Neurotoxicol. Teratol. 12:79-84; 1990. 13. Hard, E.; Musi, B.; Dahlgren, L.; Engel, J.; Larsson, K.; Liljequist, S.; Lindh, A. S. Impaired maternal behavior and altered central serotonergic activity in the adult offspring of chronically ethanol treated dams. Acta Pharmacol. Toxicol. 56:347-353; 1985. 14. Heyser, C.; Chen, W. J.; Miller, J.; Spear, N. E.; Spear, L. P. Prenatal cocaine exposure produces deficits in Pavlovian conditioning and in sensory preconditioning among infant rat pups. Behav. Neurosci., in press; 1990. 15. Hughes, J. A.; Sparber, S. B. d-Amphetamine unmasks postnatal consequences of exposure to methylmercury in utero: Methods for studying behavioral teratogenesis. Pharmacol. Biochem. Behav. 8: 365-375; 1978. 16. Isaacson, R. L.; Spear, L. P. A new perspective for the interpretation of early brain damage. In: Almli, C. R.; Finger, S., eds. Early brain damage, vol. 2. Neurobiology and behavior. New York: Academic Press; 1984:73-98. 17. Johanson, I. B.; Hall, W. G. Appetitive learning in 1-day-old rat pups. Science 205:419-421; 1979. 18. Johanson, I. B.; Hall, W. G. The ontogeny of feeding in rats: I~. Thermal determinants of early ingestive responding. J. Comp. Physiol. Psychol. 94:977-992; 1980. 19. Lanier, L. P.; Dunn, A.; Van Hartesveldt, C. The development of neurotransmitters and their function in the brain. Rev. Neurosci. 2:195-256; 1976. 20. Moody, C. A.; Frambes, N. A.; Spear, L. P. Psychopharmacological evidence for enhanced D2 dopamine receptor function following prenatal cocaine exposure. Int. Soc. Dev. Psychobiol. 30; 1989 (abstr). 21. Rajachandran, L.; Goodwin, G.; Spear, L. P. Behavioral effects of the noncompetitive NMDA antagonist MK801 in infant and preweanling rat pups. Soc. Neurosci. Abstr., in press; 1990. 22. Riley, E. P.; Hannigan, J. H.; Balaz-Hannigan, M. A. Behavioral teratology as the study of early brain damage: Considerations for the assessment of neonates. Neurobehav. Toxicol. Teratol. 7:635-638; 1985. 23. Ristine, L. A.; Spear, L. P. Effects of serotonergic and cholinergic antagonists on suckling behavior of neonatal, infant and weanling rat pups. Behav. Neural Biol. 41:99-126; 1984. 24. Rodier, P. M. Behavioral teratology. In: Wilson, J. G.; Fraser, F. C., eds. Handbook of teratology, vol. 4. New York: Plenum Press; 1978:397-428. 25. Scalzo, F. M.; Ali, S.; Frambes, N. A.; Spear, L. P. Weanling rats exposed prenatally to cocaine exhibit an increase in striatal D2 binding associated with an increase in ligand affinity. Pharmacol. Biochem. Behav. 37(2):in press; 1990. 26. Shalaby, I. A.; Spear, L. P. Psychopharmacologicai effects of low

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27. 28. 29. 30. 31.

32. 33. 34.

35. 36. 37.

38.

39.

40.

41. 42.

43.

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