Prenatal morphine exposure reduces pyramidal neurons in CA1, CA2 and CA3 subfields of mice hippocampus

Document Type: Original Article


1 Department of Anatomical Sciences, Golestan University of Medical Sciences, Gorgan, Iran

2 Gorgan Congenital Malformations Research Center, Department of Anatomical Sciences, Golestan University of Medical Sciences, Gorgan, Iran


Objective(s):This study was carried out to evaluate the effect of maternal morphine exposure during gestational and lactation period on pyramidal neurons of hippocampus in 18 and 32 day mice offspring.
Materials and Methods: Thirty female mice were randomly allocated into cases and controls. In case group, animals received morphinesulfate 10 mg/kg.body weight intraperitoneally during 7 days before mating, gestational period (GD 0-21), 18 and 32 days after delivery in the experimental groups. The control animals received an equivalent volume of normal saline. Cerebrum of six offsprings in each group was removed and stained with cresyl violet and a monoclonal antibody NeuN for immunohistochemical detection of surviving pyramidal neurons. Quantitative computer-assisted morphometric study was done on hippocampus.
Results: The number of pyramidal neurons in CA1, CA2 and CA3 in treated groups was significantly reduced in postnatal day 18 and 32 (P18, P32) compared to control groups (P<0.05).
The mean thickness of the stratum pyramidal layer was decreased in the treated groups in comparison with controls (P<0.05), whereas the mean thickness of the stratum oriens, stratum radiatum and stratum lacunosum-moleculare in CA1 field and stratum oriens, stratum lucidum, stratum radiatum and stratum lacunosum-moleculare in CA3 were significantly increased in morphine treated group in comparison with controls (P<0.05).
Conclusion: Morphine administration before and during pregnancy and during lactation period causes pyramidal neurons loss in 18 and 32 days old infant mice. 


1. Zhang Y, Chen Q, Yu LC. Morphine: a protective or destructive role in neurons? Neuroscientist 2008; 14:561-570.

2. Nestler EJ. Historical review: molecular and cellular mechanisms of opiate and cocaine addiction. Trends Pharmacol Sci 2004; 25:210-218.

3. National Institute on Drug Abuse. National Pregnancy and Health Survey: Drug use among women delivering live births: 1992 (NIH Publication 96–3819). Rockville, MD: Department of Health and Human Services 1996;1-157.

4. Ornoy A, Michailevskaya V, Lukooshov I. The developmental outcome of children born to heroin-dependent mothers, raised at home or adopted. Child Abuse Negl 1996; 20:385-396.

5. Ray JR, Dubinj W, Blechner JN. Fetal growth retardation following maternal morphine administration: nutritional or drug effect? Biol Neonal 1977; 32:222-228.

6. Wilson GS, McCreay R, Kean J, Baxter JC. The development of pre-school children of heroin-addicted mothers: a controlled study. Pediatrics 1979; 63:135-141.

7. Fazel A, Jalali M. Morphine experimentally induced exencephaly and spina bifida in mice. Arch Iran Med 2002; 5:179-183.

8. Nasiraei-Moghadam S, Sahraei H, Bahadoran H, Sadooghi M, Salimi SH, Kaka GR,et al. Effects of maternal oral morphine consumption on neural tube development in Wistar rats. Brain Res Dev Brain Res 2005; 159:12-17.

9. Chooa RE, Huestis MA, Schroeder JR, Shin AS, Jones HE. Neonatal abstinence syndrome in methadone-exposed infants is altered by level of prenatal tobacco exposure. Drug Alcohol Depend 2004; 75:253-260.

10. Mao J, Sung B, Ji RR, Lim G. Neuronal apoptosis associated with morphine tolerance: evidence for an opioid-induced neurotoxic mechanism. J Neurosci 2002; 22:7650-7661.

11. Atici S, Cinel L, Cinel I, Doruk N, Aktekin M, Akca A,et al. Opioid neurotoxicity: comparison of morphine and tramadol in an experimental rat model. Int J Neurosci 2004; 114:1001–1011.

12. Turchan-Cholewo J, Liu Y, Gartner S, Reid R, Jie C, Peng X,et al. Increased vulnerability of ApoE4 neurons to HIV proteins and opiates: protection by diosgenin and L-deprenyl. Neurobiol Dis 2006; 23:109–119.

13. Bekheet SH, Saker SA, Abdel-Kader AM, Younis AEA. Histopathological and biochemical changes of morphine sulphate administration on the cerebellum of albino rats. Tissue Cell 2010; 42:165–175.

14. Ghafari S, Roshandel D, Golalipour MJ. Effect of intrauterine morphine sulfate exposure on cerebellar histomorphological changes in neonatal mice. Folia Neuropathol 2011; 49:328-334.

15. Svensson AL, Bucht N, Hallberg M, Nyberg F. Reversal of opiate-induced apoptosis by human recombinant growth hormone in murine foetus primary hippocampal neuronal cell cultures. Proc Natl Acad Sci USA 2008; 105:7304-7308.

16. Mei B, Niu L, Cao B, Huang D, Zhou Y. Prenatal morphine exposure alters the layer II/III pyramidal neurons morphology in lateral secondary visual cortex of juvenile rats. Synapse 2009; 63:1154-1161.

17. Seatriz JV, Hammer RP Jr. Effects of opiates on neuronal development in the rat cerebral cortex. Brain Res Bull 1993; 30:523-527.

18. Eisch AJ, Barrot M, Schad CA, Self D, Nestler EJ. Opiates inhibit neurogenesis in the adultrat hippocampus. Proc Natl Acad Sci USA 2000; 97:7579–7584.

19. Niu L, Cao B, Zhu H, Mei B, Wang M, Yang Y, Zhou Y. Impairedin vivo synaptic plasticity in dentate gyrus and spatial memory in juvenile rats induced by prenatal morphine exposure. Hippocampus 2009; 619:649-657.

20. Emeterio EP, Tramullas M, Hurlé MA. Modulation of apoptosis in the mouse brain after morphine treatments and morphine withdrawal. J Neurosci Res 2006; 83:1352-1361.

21. Hauser KF, Gurwell JA, Turbek CS. Morphine inhibits Purkinje cell survival and dendritic differentiation in organotypic cultures of the mouse cerebellum. Exp Neurol 1994; 130:95–105.

22. Hauser KF, Harris-white ME, Jackson JA, Opanashuk LA, Carney JM. Opioids disrupt Ca2+ homeostasis and induce carbonyl oxyradical production in mouse astrocytes in vitro: transient increases and adaptation to sustained exposure. Exp Neurol 1998; 151:70-76.

23. Hauser KF, Houdi AA, Turbeck CS, Elde RP, Maxson W. Opioids intrinsically inhibit the genesis of mouse cerebellar granule neuron precursorsin vitro: differential impact of mu and delta receptor activation on proliferation and neurite elongation. Eur J Neurosci 2000; 12:1281–1293.

24. Hauser KF, McLaughlin PJ, Zagon IS . Endogenous opioids regulate dendritic growth and spine formation in developing rat brain. Brain Res 1987; 416:157-161.

25. Lorber BA, Freitag SK, Bartolome JV. Effects of beta-endorphin on DNA synthesis in brain regions of preweanling rats. Brain Res 1990; 531:329-332.

26. Schmahl W, Funk R, Miaskowski U, Plendl J. Long-lasting effects of naltrexone, an opioid receptor antagonist, on cell proliferation in developing rat forebrain. Brain Res 1989; 486:297-300.

27. Hammer RPJ, Ricalde AA, Seatriz JV. Effects of opiates on brain development. Neurotoxicology 1989; 10:475-483.

28. Zagon IS, McLaughlin PJ. Endogenous opioids regulate cell proliferation in the developing rat brain. Brain Res 1987; 412:68-72.

29. Zagon IS, Mc Laughlin PJ. Identification of opioid peptides regulating proliferation of neurons and glia in the developing nervous system. Brain Res 1991; 542:318–323.

30. Hauser KF, Osborne JG, Stiene-Martin A, Melner MH. Cellular localization of proenkephalin mRNA and enkephalinpeptide products in cultured astrocytes. Brain Res 1990; 522:347-353.

31. Spruce BA, Curtis R,Wilkin GP, Glover DM. Aneuropeptide precursor in cerebellum: Proenkephalin exists in subpopulations of both neurons and astrocytes. EMBO J 1990; 9:1787–1795.

32. Shinoda H, Marini AM, Cosi C, Schwartz JP. Brain region and gene specificity of neuropeptide gene expression in cultured astrocytes. Science 1989; 245:415-417.

33. Zhu Y, Hsu MS, Pintar JE. Developmental expression of the Mu, kappa and delta opioid receptor mRNAs in mouse. J Neurosci 1998; 18:2538-2549.

34. Villarreal DM, Derrick B, Vathy I. Prenatal morphine exposure attenuates the maintenance of late LTP in lateral perforant path projections to the dentate gyrus and the CA3 regionin vivo. J Neurophysiol 2008; 99:1235-1242.

35. Cheng