In humans, lack of phenylalanine hydroxylase (Pah) activity results in phenylketonuria (PKU), which is associated with the development of severe mental retardation after birth. The underlying mechanisms, however, are poorly understood. Mutations of the Pah gene in Pah(enu2)/c57bl6 mice result in elevated levels of phenylalanine in serum similar to those in humans suffering from PKU. In our study, long-term potentiation (LTP) and paired-pulse facilitation, measured at CA3-CA1 Schaffer collateral synapses, were impaired in acute hippocampal slices of Pah(enu2)/c57bl6 mice. In addition, we found reduced expression of presynaptic proteins, such as synaptophysin and the synaptosomal-associated protein 25 (SNAP-25), and enhanced expression of postsynaptic marker proteins, such as synaptopodin and spinophilin. Stereological counting of spine synapses at the ultrastructural level revealed higher synaptic density in the hippocampus, commencing at 3 weeks and persisting up to 12 weeks after birth. Consistent effects were seen in response to phenylalanine treatment in cultures of dissociated hippocampal neurones. Most importantly, in the hippocampus of Pah(enu2)/c57bl6 mice, we found a significant reduction in microglia activity. Reorganization of hippocampal circuitry after birth, namely synaptic pruning, relies on elimination of weak synapses by activated microglia in response to neuronal activity. Hence, our data strongly suggest that reduced microglial activity in response to impaired synaptic transmission affects physiological postnatal remodelling of synapses in the hippocampus and may trigger the development of mental retardation in PKU patients after birth.
A well-balanced activity of the aryl hydrocarbon receptor (AhR) is necessary for normal ovarian function. As known from murine AhR knock-out (KO) models, the AhR is involved in folliculogenesis, gonadotrophin receptor expression, proliferation of granulosa cells and intraovarian estrogen signalling. Highly potent, non-physiological ligands such as the dioxin 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) lead to a blockade of ovulation, estrogen receptor degradation and reduction of estrogen levels. Estrogen synthesis is a typical function of granulosa cells and essential for normal cyclicity and fertility. We employed the human granulosa cell line KGN to further characterize AhR signalling and AhR function in granulosa cell physiology. Real-time PCR quantification of the target genes Cyp1a1 and Cyp1b1 and reporter gene assays after stimulation with TCDD or beta-naphthoflavone (BNF) or inhibition with alpha-naphthoflavone (ANF) or 3'-methoxy-4'-nitroflavone (3,4-MNF) of the AhR demonstrated constitutive activity and functionality of AhR pathway in KGN granulosa cells. In untreated KGN cells, AhR protein was exclusively detected in the nuclear fraction. TCDD stimulation affected the gonadotrophin receptor but not estrogen receptor β (ERβ) protein expression. Additionally, the constitutively activated AhR suppressed aromatase expression and estrogen synthesis (enzyme-linked immunoassay, ELISA) and enhanced proliferation [Bromodeoxyuridine (BrdU) ELISA] of KGN cells. Activation of the AhR by BNF did not override this inhibitory effect on estrogen synthesis or proliferation. In conclusion, the AhR pathway is constitutively activated and functional in human KGN granulosa cells. It is a potential target for endocrine disruption by exogenous ligands and subsequent dysfunction of granulosa cells.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.