Long-term alterations in brain function following cocaine administration during the preweanling period

309

Del'elopmental Brain Research, 72 (1993) 309-313 (~> 1993 Elsevier Science Publishers B.V. All rights reserved (1165-3806/93/$06.0(}

BRESD 60488

Long-term alterations in brain function following cocaine administration during the preweanling period D i a n a L. D o w - E d w a r d s ,

Laurel A. Freed-Malen

a n d H a r r y E. H u g h e s

Laboratot3' o1' Cerebral Metabolism, Department of Pharmacolo~3'. State Unirersity qf New York. Health Science Center, Brook(vn, NY 112(I.? (USA; (Accepted 22 December 1992)

Key words: Development; Stimulant; Glucose metabolism; Long-term effect

This report examines the long-term effects of cocaine exposure during postnatal days (P) 11-20 on the metabolic function of major central neuronal systems. Cocaine (50mg/kg) or vehicle was administered subcutaneously to rat pups during P 11-20. At 60 64 days of age, the rats were examined for cerebral glucose metabolic patterns. In cocaine-treated females 18 of the 46 structures evaluated showed increased metabolic rates including 5 of 6 structures within the motor system and 7 of 17 limbic structures. No decreased rates were seen. In malcs, cocaine had no effects in the motor structures or hypothalamus while 2 of 17 structures within the limbic system showed decreased rates of glucose utilization and 2 of I1 structures within the sensory systems showed increased rates. These results indicate that female rats show greater long-term metabolic effects than males and that cocaine exposure during P 11-2{) produces different metabolic effects than cocaine exposure during P I-10 which we previously reported.

Although cocaine use has decreased nationwide within the last few years, a significant number of infants and children have been exposed to the drug (see ref. 9 for review). Exposed infants have been described as having a range of symptoms from mild and temporary neurobehavioral abnormalities to severe growth and mental retardation (see ref. 3 for review). Identifying the biological effects of a drug in a clinical population can be difficult particularly when the drug is used in concert with several other drugs and by individuals with multiple risk factors. Researchers have turned to animal models in an effort to determine what effects cocaine may have on the development of central neuronal systems. During the past few years, several groups have identified neurobehavioral abnormalities in preweanling rats exposed to cocaine during prenatal life l'L~'14 and we have identified brain metabolic effects in adult rats exposed to cocaine either prenatally or during the early postnatal period a'5. Our studies showed that cocaine exposure during prenatal development reduced glucose metabolic activity in several brain regions including the

hypothalamus, the nigrostriatal pathway and in specific structures within the limbic system in male rats 4. Cocaine administration during postnatal days (P) 1-10, however, had no long-term effect in male rats but produced significant increases in metabolism primarily in the limbic system in female rats 5. In order to determine what effect cocaine might have on development of the brain during P 11-20, a time when forebrain, hypothalamus, and cortex are undergoing synaptic maturation, we administered cocaine during this period and examined adult rats using the quantified deoxyglucose method of Sokoloff as an indicator of brain function ~2. Some of these data have been presented in abstract form =. Sprague-Dawley rats (Charles River, Wilmington, ME) were mated in our AALAC-accredited animal facility and left undisturbed until the day of birth. Litters were culled to eight with equal numbers of males and females. Beginning on P 11 (day of birth was P 1), half the pups received 50 m g / k g / 5 ml cocaineHC1 (Sigma Chemical Co.) and the other half received 5 m l / k g of the vehicle (sterile water) subcutaneously.

Corre,~pondence." D. Dow-Edwards, Department of Pharmacology, Box 29, State University of New York, 45(l ('larkson Ave.. Brooklyn, NY 112113, USA. Fax: (1)(718)270-2241.

311) Daily injections continued up to and including P20. Pups were weaned on day 21 into cages containing same gender littermates and weighed every 4 days until P60. At 60 days of age, the quantified deoxyglucose method of measuring brain glucose metabolism le was carried out on males and diestrus females as determined by daily vaginal smears from postnatal day 56 until a diestrus smear was seen after a full cycle was observed. Following an overnight fast, arterial and venous catheters were inserted and passed subdermally to the back of the neck while the animal was under halothane anesthesia. Following a 2 h recovery period, during which the body t e m p e r a t u r e was maintained at the preanesthetic temperature, each rat was administered 125 /~Ci/kg body weight of [~4C]2-deoxy-m-glucose (spec. act. 55 m C i / m M , American Radiolabelled Corp., St. Louis, MO) and timed arterial samples taken for the determination of plasma 14C and glucose levels using the Beckman LS 1800 scintillation counter and YSI 23A glucose analyzer respectively. During the period of isotope circulation, the rats were freely moving in 18 ( W ) x 29 ( L ) x 11 (H) cm Plexiglas cages with shavings and only had water available to drink. Since the cages were placed on the lab bench, the animals were exposed to the typical lab environment with the exception that effort was made to minimize extraneous noise. In all cases, the period of isotope circulation was between noon and 3PM. Following 45 min. of isotope circulation, the animal was administered a lethal dose of sodium pentobarbital (60 rag). The brain was removed, frozen and later sectioned at 20 /xm for deoxyglucose autoradiography and histology. Sections were placed against Kodak OM film along with calibrated ~4C standards for 12 days. Following film development, brain regions of interest were evaluated using

the Loats or M C I D imaging systems by individuals unaware of the experimental condition of the animals. Structures were identified based on the atlas of Paxinos and Watson I1 and alternate thionin-stained sections. At least 6 sections per structure per animal were evaluated. Statistical comparisons between the two groups of animals were made using the Student's t-test with P < 0.05 used as the acceptable level of significance. Males and females were evaluated independently. Since a total of 108 t-tests were performed in this study, one should expect to find statistically significant effects in up to 5 comparisons due to chance alone; the type I statistical error. In addition, statistically significant differences in about 20% of the comparisons may have been missed due to type II statistical errors. Cocaine administration during P l l - 2 0 decreased body weight gain during the period of drug administration. For example, control rats gained 21 +_ 0.6 g (mean + S.E.M.) between P 11 and P 20 while cocaine-treated rats gained only 15 + 0.6 g. However, once drug administration was terminated, the cocaine-treated rats quickly gained weight and at 60 days of age were not significantly different from controls (Table I). Cocaine exposure during P 11-20 resulted in widespread increases in glucose metabolic activity in female rats compared to controls and selected increases and decreases in males (Table II). In the females, cortex showed the greatest concentration of effects with all but the auditory and parietal cortices showing statistically significant increases in metabolism. Five of six structures associated with the motor system also showed increases in metabolism including the forelimb motor cortex, the caudate, the substantia nigra pars compacta, the substantia nigra pars reticulata, and the red nucleus. On the other hand, the limbic

TABLE 1 Physiologic parameters of rats treated postnatally (days 11-20) with cocaine

Values are means-+S.E.M. for the number of animals indicated. Females

Body weight (g) Glucose (mg/ml) Temperature(°C) Arterial pressure (mmHg) Hematocrit (%) Arterial blood gases PO2(mmHg) pCO2(mmHg) pH Estrus cycle

Males

Water n=7

Cocaine n=5

239 + 5 1.04 ± 0.05 37.9 ±0.3 114 ±2 44.6 ±3.2

250 + 10 0.97 ± 0.05 37.9 ± 0.2 116 ± 2 48.2 ± 0.7

84.9 ±3.1 29.9 +0.9 7.39±0.01 diestrus

84.1 +_ 5.1 28.7 -+ 0.6 7.39± 0.01 diestrus

Water

n=7 395 + 8 0.93 + 0.04 37.4 ___0.1 114 ±3 46.2 ± 1.2 77.4 ±4.7 31.7 -+0.9 7.43±0.01

Cocaine n=8

394 ± 7 0.97 ± 0.03 37.4 ±0.1 110 _+1 47.[) ± 1.4 82.7 ±3.4 30.3 ±1.3 7.44±0.01

311 T A B L E 1I Rates ~f glucose metabolism in selected brain regions from rats treated with cocaine during postnatal days 11- 20

Data are in # m o l / 1 0 0 g tissue/rain; mean_+ S.E.M. for the numbers of animals indicated, Female rats Control (n = 5)

Motor structures motor cx/fl~relimb caudaten. globuspallidus substantia nigra reticulata compacta redn. Limbic structures Mesolimbic forebrain accumbens vert. limbdiag, band septumlateral Amygdala lateral central cortical Limbic cortex cingulate piriform Hippocampus CAI molecular layer dentate gyrus Habenula media/ lateral ventraltegmentalarea interpeduncularn. medialraphen. thalamus mediodorsal Hypothalamic structures mammillary body suprachiasmaticn arcuaten median eminence ventromedialn dorsomedialn lateral paraventricular n Sensory structures sensorycx/forelimb ventralthalamicn. vestibularn. occipitalcx. lateralgeniculaten. superior colliculus auditorycx. medialgeniculaten. inferior colliculus vent. caud. lat. lemnis superior olive Association areas parietal cortex zonaincerta p o n t i n e r e t , form.

Male rats

~A

Cocaine (n = 5)

%A

Control (n = 7)

Co~'ame (n = 6)

92+_ 4 94_+ 4 53+ 4

108+_ 6 " 114_+ 8 " 59+ 2

+18 +22 +11

122+7 124+7 62_+3

117+ 6 117+ 4 64_+ 2

7 -7 +3

57_+ 3 71+_ 4 86+_ 4

67_+ 3 * 85+ 4* 100_+ 5 *

+18 +19 +17

73_+3 89+3 101+5

71+ 3 85+ 2 103+ 4

+1 -3 +2

77-+ 6 75-+ 6 51-+ 3

87+ 7 96-2- 5 " 69_+ 5

+12 +28 +18

94+6 91 + 5 60+2

76+ 3 * 94+ 4 58+ 1

73-+ 5 38_+ 2 66 + 5

94_+ 6 " 46_+ 4 85 + 8

+28 +21 + 29

95+4 43+2 89-+ 4

95+ 6 43+ 2 88-+ 4

99_+ 5 121_+ 6

124_+10" 142-+ 6 *

+25 +18

136+9 149-+5

128+ 6 135_+ 4 *

67_+ 4 86-+ 4 52-+ 3

82_+ 6 100+ 5 70 ± 6 *

+21 +16 + 34

96+4 115_+5 68 + 3

88+_ 3 109+ 2 66 +_ 2

8 5 - 3

-2(I +3 3 11 11 l -8 -12

63+ 101+ 63_+ 107_+ 90+ 116_+

4 7 4 7 8 7

91_+1{1" 133_+11 * 71/_+ 1 124_+ 5 106_+ 6 137_+ 8

+44 +32 +11 +16 +18 + 18

76_+4 132_+7 72_+5 119_+5 102+2 145_+7

73+ 123+ 75+ 124+ 106-+ 145+

3 7 2 6 3 5

-4 -7 +4 +4 +4 11

120_+ 58_+ 35+ 44_+ 40_+ 51+ 55+ 55-+

3 4 4 4 4 4 3 3

128+ 10 72-+ 6 58-+11/* 51+ 7 40+ 4 58_+ 4 63-+ 3 68_+ 2 *

+6 +24 +65 +16 0 +14 +15 + 23

137_+4 71-+2 57_+3 50_+4 44_+3 56+_2 70+3 56_+ 3

137+ 10 73+ 4 47_+ 4 49+ 6 511+ 3 62+ 3 7l_+ 3 67 + 5

0 +3 +12 +4 ~14 +11 +3 +2

89_+ 90_+ 131_+ 96_+ 87+_ 94_+

4 5 7 6 6 4

110_+ 108+ 128-+ 112-+ 104_+ 98_+

+24 +20 -3 +17 +20 +4

120_+6 121_+5 134+_6 126+_6 112_+4 105-+3

115+ 122+ 147+ 119+ 113_+ 1111+

6 5 6 3 3 4

7 +3 +10 b~ 0 +6

+8 +4 0 0 -4

141+5 139+4 169+7 121 + 4 135_+4

161-+ 154-+ 172_+ 126+ 138+

7 * 6" 7 5 9

+21 +17 +6 +7 +7

+19 +25 +17

118+6 91_+6 77+3

1111+ 5 96+ 5 79+ 3

131-+ 6 123+ 4 160+_ 10 126-+ 7 150+13 93+ 7 76+ 3 65-+ 7

6" 3" 6 4* 5 4

142_+10 128_+10 160_+ 8 126+_ 3 144_+ 8 111+ 6 95_+ 3 * 76_+ 4

* Significant difference from control value, P < 0.05 by t-test.

-7 +5 +3

312 system, hypothalamus, sensory systems and association areas showed more selective increases in metabolism. Within the subcortical limbic system, the lateral septurn, the vertical limb of the diagonal band, the lateral amygdala, the dentate gyrus and the medial and lateral habenula showed increases. The arcuate and paraventricular nuclei of the hypothalamus also showed significant increases as did the ventral thalamic nucleus and the zona incerta. In males, only two structures showed increases, the auditory cortex and the medial geniculate body, while two showed decreases, the n. accumbens and piriform cortex. Alterations in glucose utilization were not associated with alterations in any of the physiologic parameters assessed including basal plasma glucose concentration, mean arterial blood pressure, blood gases or pH, or hematocrit (Table I). Administration of cocaine at 50 m g / k g to rat pups during P 11-20 produced long-term changes in glucose metabolism in many brain regions in females and highly localized changes in males. Although this gender difference resembles that observed following cocaine administration during P 1-105, the neuronal systems affected differ following the two exposure periods. That is, whereas cocaine exposure during P I - 1 0 resulted in increased glucose metabolism in the medial portions of the mesencephalic dopamine system and other selected structures within the limbic system in female rats, exposure during P l l - 2 0 resulted in increased metabolism in the lateral portions of the mesencephalic dopamine system and the cortex in general. Thus, exposure during P I 1 - 2 0 produced increased metabolism in the substantia nigra and caudate nucleus and not the ventral tegmental area or the nucleus accumbens. Exposure during P I - 1 0 produced increased metabolism in several midline nuclei within the limbic system including the interpeduncular nucleus and medial raphe; exposure during P 11-20 did not. The medial and lateral habenula were the only subcortical structures within the limbic system showing increased metabolism following both exposure periods. Cocaine exposure during P l l - 2 0 affected cortical metabolism more so than other brain regions in female rats; i.e., five of seven cortical structures analyzed showed increased rates of metabolism. There is ample evidence that maturation of pre and postsynaptic elements is occurring in the cortex during this time and that manipulation of neurotransmitter systems can alter this process (see ref. 10 for review). Since cocaine, through inhibition of reuptake, enhances activity in at least threc neurotransmitter systems in adult brain; dopamine, serotonin and norephinephrine; the pres-

ence of cocaine during this critical period ot development may alter the process of synaptic maturation in any or all of these neurotransmitter systems. The effects of cocaine on cerebral development cannot be explained solely on the basis of the drug interacting with the maturation of synapses. That is, while synapses most likely mature with a similar time course in male and female rats, effects of cocaine were seen primarily in female rats. In male rats, cocaine exposure during P l l - 2 { ) resulted in increased metabolism in only two structures and decreases in two other structures. The reason for the striking gender difference is not immediately clear. Preliminary data in our laboratory indicate that plasma cocaine levels in female rat pups are similar to those in males following 50 m g / k g cocaine. However, differences in neuroendocrine systems as well as neurotransmitter systems, including the serotonin system have been identified in male and female preweanling rats 6'~. In addition, male rats are not entirely protected from the developmental effects of cocaine since prenatal cocaine exposure resulted in adult male offspring with decreases in glucose utilization in primary motor and somatosensory cortices, several subcortical dopaminergic structures as well as the majority of the hypothalamic structures evaluated 4. Therefore, a detailed analysis of the interaction of cocaine with each of these variables, i.e., gender differences in neuroendocrine function, brain maturation and metabolism, will be necessary before definitive conclusions regarding the striking gender differences in response to cocaine can be drawn. These studies were supported by the National Institute on Drug Abuse Grant DA04118. 1 Church, M.W. and Overbeck, G.W., Prenatal cocaine exposure in the Long-Evans rat: II. Dose-dependent effects on offspring behavior, Neurotoxieol. Teratol., 12 (1990)335-343. 2 Dow-Edwards, D.L., Functional effects of cocaine given during critical periods of development, J. Cerebral Blood Flow Metab., 9 (suppl. 1) (1989) $712 (abstr.). 3 Dow-Edwards, D.L., Cocaine effects on fetal development: A comparison of clinical and animal research findings, NeurotoxicoL TeratoL, 13 (1991) 347-352. 4 Dow-Edwards, D.L., Freed, L.A. and Fico, T.A., Structural and functional effects of prenatal cocaine exposure in adult rat brain, Dev. Brain Res., 57 (1990) 263-268. 5 Dow-Edwards,D.L., Freed, L.A. and Milhorat, T.H., Stimulation of brain metabolism by perinatal cocaine exposure, Det,. Brain Res., 42 (1988) 137-14l. 6 Giulian, D., Pohorecky, L.A. and McEwen, B.S., Effects of gonadal steroids on brain 5-hydroxytryptaminelevels in the neonatal rat, Endocrinology, 93 (1973) 1329-1335. 7 Hutchings, D., Fico, T.A. and Dow-Edwards, D.L., Prenatal Cocaine: maternal toxicity, fetal effects and locomotor activity in rat offspring, Neurotoxicol. TeratoL, 11 (1989)65-69. 8 Ladosky,W. and Gaziri, L.C.J., Brain serotonin and sexual differentiation of the nervous system, Neuroendocrinology, 6 (1970) 168-174.

313 9 Neuspiel, D.R. and Hamel, S.C., Cocaine and infant behavior, Def. Behal'. Pediatr., 12 (1991) 55-64. 10 Parnavelas. J.G., Papadopoulos, G.C. and Cavanagh, M.E., Changes in neurotransmitters during development. In A. Peters and E.G. Jones (Eds.), Cerebral Cortcv, Vol. 7, Plenum. New York, 198% pp. 177-210. 11 Paxinos, G. and Watson, C., The Rat Brain in Stereotaxic Coordinates, Academic Press, New York, 1982. 12 Sokoloff, 1.., Reivich, M., Kennedy, C., DesRosiers, M.tt., Patlak, C.S., Pettigrew, K.D., Sakurada, O. and Shinohara, M., The [14C]deox3'glucose method for the measurement of local cerebral glucose utilization: theory., procedure, and normal values in the

conscious and anesthetized albino rat, J. Neuroc/wm., 28 (1977) 897-916. 13 Spear, L.P., Kirstein, C.L., Bell, J., Yoottanasumpun, V., Grcenbaum, R., O'Shea, J., Hoffman H. and Spear. N.E., L~ffects of prenatal cocaine exposure on behavior during the early poslnatal period, NeurotoxicoL Yeratol., I1 (1989a)57 63. 14 Spear, L.P., Kirstcin, C.L. and Frambes, N.A., Cocaine effects on developing central nervous system: behavioral, psychopharmacological, and neurochemical studies. In D.E. Hutchings (Ed.), U.w of l.icit and l/licit Drugs During Pre~nam'y, Ann. N'e~ Y~rk Acad. Sci.. VoL 562, New York. NY, 19~9. pp. 290 307.