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Friday, May 8, 2009: 2:10 PM
Northwest Hall Room 1 (Chicago Hilton)
Background:
Reelin is a candidate gene for neurodevelopmental disorders such as autism and schizophrenia. Reelin haploinsufficiency in the heterozygous rl/+ mouse causes behavioral abnormalities soon after birth, consisting of decreased ultrasound vocalizations (USV) emitted by pups upon maternal separation, as well as in adult life, producing behavioral rigidity in a task requiring a change in strategy. At the anatomical level, reelin haploinsufficiency causes a loss of GABAergic inhibitory neurons, e.g. parvalbumin-positive neurons in limbic areas and basal ganglia, and Purkinje cells (PC) in the cerebellum. This PC loss is more evident in male than female mice.
Objectives:
1) To characterize brain circuit abnormalities of heterozygous reeler (rl/+) mice, and their sex-dependency; 2) To unravel the mechanisms of neuronal loss in brain areas that are relevant for autism, like the cerebellum and limbic system, focusing on interactions between decreased reelin levels and sex steroids; 3) To characterize the relationship between circuit alterations and behavioral abnormalities of rl/+ mice, in particular decreased USV.
Methods:
1) Assessment of neuronal numbers with unbiased stereology; 2) Manipulation of brain levels of sex steroids using agonists and antagonists; 3) Measurement of reelin mRNA levels with RT-PCR; 4) Assessment of USV emitted by pups after separation from the mother; 5) Assessment of behavioral flexibility in a strategy-changing task (Intradimensional-Extradimensional Set Shifting Task).
Results:
PC numbers are selectively reduced in rl/+ males at postnatal day 15, with no parallel loss of their main presynaptic input neurons, i.e. granule cells and inferior olivary neurons. Early postnatal administration of the estrogen receptor (ER) agonist 17β-Estradiol (17βE) in the cisterna magna leads to recovery of PC numbers in male rl/+, but has no effect in female rl/+, or +/+ mice of either sex; conversely, ER antagonists 4-OH-Tamoxifen or ICI 182,780 selectively reduce PC numbers in female +/+ and rl/+, while the same ER antagonists have no effect in male rl/+ or +/+ mice. RT-PCR analysis revealed that 17βE administration restores reelin mRNA levels in rl/+ mice, indicating that transcription of the reelin gene is under 17βE control in the cerebellum. Administration of 17βE in the cisterna magna at P5, in doses leading to recovery of PCs, also leads to an increase of USV of rl/+ mice, in response to maternal separation, as well as a better test performance in the adult rule-shifting task.
Conclusions:
Taken together, these results strongly suggest that reelin and 17βE interact during neurodevelopment to sculpt cerebellar circuitry. This effect may represent a novel model to understand how a genetic mutation interacts with perinatal sex hormones, leading to gender-dependent abnormalities of neural circuits and behavior that persist into adulthood. Furthermore, in conjunction with recent experimental evidence showing that brain levels of 17βE are rapidly modulated by the social context, similar to other classical neuromodulators, our results suggest a novel way of understanding mechanisms by which the environment permanently sculpts neural circuits during brain development. Finally, our data may help to build a neurobiological foundation for the extreme male brain theory of autism.
Reelin is a candidate gene for neurodevelopmental disorders such as autism and schizophrenia. Reelin haploinsufficiency in the heterozygous rl/+ mouse causes behavioral abnormalities soon after birth, consisting of decreased ultrasound vocalizations (USV) emitted by pups upon maternal separation, as well as in adult life, producing behavioral rigidity in a task requiring a change in strategy. At the anatomical level, reelin haploinsufficiency causes a loss of GABAergic inhibitory neurons, e.g. parvalbumin-positive neurons in limbic areas and basal ganglia, and Purkinje cells (PC) in the cerebellum. This PC loss is more evident in male than female mice.
Objectives:
1) To characterize brain circuit abnormalities of heterozygous reeler (rl/+) mice, and their sex-dependency; 2) To unravel the mechanisms of neuronal loss in brain areas that are relevant for autism, like the cerebellum and limbic system, focusing on interactions between decreased reelin levels and sex steroids; 3) To characterize the relationship between circuit alterations and behavioral abnormalities of rl/+ mice, in particular decreased USV.
Methods:
1) Assessment of neuronal numbers with unbiased stereology; 2) Manipulation of brain levels of sex steroids using agonists and antagonists; 3) Measurement of reelin mRNA levels with RT-PCR; 4) Assessment of USV emitted by pups after separation from the mother; 5) Assessment of behavioral flexibility in a strategy-changing task (Intradimensional-Extradimensional Set Shifting Task).
Results:
PC numbers are selectively reduced in rl/+ males at postnatal day 15, with no parallel loss of their main presynaptic input neurons, i.e. granule cells and inferior olivary neurons. Early postnatal administration of the estrogen receptor (ER) agonist 17β-Estradiol (17βE) in the cisterna magna leads to recovery of PC numbers in male rl/+, but has no effect in female rl/+, or +/+ mice of either sex; conversely, ER antagonists 4-OH-Tamoxifen or ICI 182,780 selectively reduce PC numbers in female +/+ and rl/+, while the same ER antagonists have no effect in male rl/+ or +/+ mice. RT-PCR analysis revealed that 17βE administration restores reelin mRNA levels in rl/+ mice, indicating that transcription of the reelin gene is under 17βE control in the cerebellum. Administration of 17βE in the cisterna magna at P5, in doses leading to recovery of PCs, also leads to an increase of USV of rl/+ mice, in response to maternal separation, as well as a better test performance in the adult rule-shifting task.
Conclusions:
Taken together, these results strongly suggest that reelin and 17βE interact during neurodevelopment to sculpt cerebellar circuitry. This effect may represent a novel model to understand how a genetic mutation interacts with perinatal sex hormones, leading to gender-dependent abnormalities of neural circuits and behavior that persist into adulthood. Furthermore, in conjunction with recent experimental evidence showing that brain levels of 17βE are rapidly modulated by the social context, similar to other classical neuromodulators, our results suggest a novel way of understanding mechanisms by which the environment permanently sculpts neural circuits during brain development. Finally, our data may help to build a neurobiological foundation for the extreme male brain theory of autism.