Klotho regulates CA1 hippocampal synaptic plasticity
Introduction
The klotho (KL) protein was originally characterized for its lifespan effects (Kuro-o et al., 1997, Kurosu et al., 2005, Dubal et al., 2014). Overexpression of KL is sufficient to extend mouse lifespan (Kurosu et al., 2005, Dubal et al., 2014). In contrast, KL-deficiency shortens mouse lifespan to ∼8 weeks by disrupting phosphate homeostasis (Kuro-o et al., 1997). Increasing evidence implicates a brain-specific function for KL. Short lived KL-deficient (KO) mice rapidly develop cognitive impairment between the 5th and 7th week of life (Nagai et al., 2003). Meanwhile long-lived KL overexpressing (OE) mice show increased cognitive function after ∼6 months of age (Dubal et al., 2014). As well, KL overexpression is sufficient to protect against the development of Alzheimer’s disease-like cognitive and synaptic impairment (Dubal et al., 2015) and may be neuroprotective (Zeldich et al., 2014). Cognitive impacts of KL extend to humans, as those carrying the functional KL-VS polymorphism have increased circulating KL (Arking et al., 2002), larger prefrontal cortical volume, and enhanced cognitive function (Dubal et al., 2014, Yokoyama et al., 2015).
The profound and inverse effects of KL on cognition are not easily explained by the data collected to date which broadly characterize the KO and OE brain. KO brains show slightly increased levels of apoptotic (Shiozaki et al., 2008) and oxidative stress (Nagai et al., 2003) proteins, mild decreases of some synaptic proteins (Shiozaki et al., 2008), a slight stochastic increase in neuron death (Nagai et al., 2003), and loss of some myelinated fibers and oligodendrocytes (Chen et al., 2013). Although OE mouse brains show no cellular changes, neurons are more resistant to oxidative stress (Brobey et al., 2015) and hippocampal NMDA GluN2B receptor subunit expression is upregulated (Dubal et al., 2014). At the level of synaptic function, long-term potentiation (LTP) of KO CA1 neurons is mildly decreased but only when lightly stimulated by a single theta burst (Park et al., 2013). Meanwhile, adult OE hippocampal dentate gyrus recordings show increased LTP (Dubal et al., 2014).
KL is expressed by both neurons and the choroid plexus, the endothelial cells that form the blood:cerebrospinal fluid barrier and generate cerebrospinal fluid (Li et al., 2004, Clinton et al., 2013). The mouse brain is the only organ where all forms of KL are detected: transmembrane, shed, and secreted (Imura et al., 2004, Clinton et al., 2013, Masso et al., 2015). Peripheral systems that characterized KL’s functions showed that KL protein function varies with its form. Transmembrane KL is a co-receptor that transduces signaling of fibroblast growth factor 23 (FGF23) (Kurosu et al., 2006). Transmembrane KL is shed from cell surfaces into serum and cerebrospinal fluid (Imura et al., 2004, Chen et al., 2007, Yokoyama et al., 2015). Shed KL is acts as a sialidase (Cha et al., 2008, Cha et al., 2009) and a signaling pathway inhibitor (Kurosu et al., 2005, Liu et al., 2007, Doi et al., 2011, Zhou et al., 2013). The function of secreted KL remains unknown (Masso et al., 2015).
Little is known about the brain-specific mechanisms of KL action. We sought to determine whether KL is detected at the synapse, and found that transmembrane KL is a synaptic protein which could function to directly affect synaptic transmission. Thus we measured synaptic transmission before and after the onset KO and OE cognitive changes (Nagai et al., 2003, Dubal et al., 2014). We observed KL-mediated effects on both pre- and post-synaptic function of hippocampal CA1 Schaffer collateral neurons and attempted to rescue synaptic plasticity with acute application of shed KL.
Section snippets
Animals
KL-deficient (KO,129S1/SvImJ) and overexpressing (OE,C57BL/6J) lines of mice are from M. Kuro-o (University of Texas Southwestern and Jichi Medical University, Japan). Strain-specific, wild-type (WT) controls were used for all experiments. All mice were housed with free access to food and water at 26.6 °C, humidity maintained above 40%. KO mice were supplemented with Bacon Softies or Nutra-gel (BioServ, Frenchtown, NJ) beginning week 5. All procedures were approved by the University of Alabama
Synaptic membranes contain Klotho
First, we confirmed that KL is generated by multiple cell types within the brain of WT but not KO mice (Fig. 1). Consistent with previous reports (Li et al., 2004, Maltare et al., 2014), choroid plexus KL immunoreactivity is most intense (Fig. 1A). Although less highly expressed than choroid plexus, throughout subfields of the hippocampus, neuronal cell bodies and processes express KL (Fig. 1B). CA1 neurons prominently show that KL co-localizes with dendritic marker Map2 but not axonal marker
Discussion
Our data show that hippocampal neurons express KL and synaptic membranes contain the full length protein (Fig. 1, Fig. 2). KL-deficiency enhances pre- and post-synaptic plasticity of Schaffer collateral neurons prior to the onset of cognitive impairment but without affecting initial release probability or NMDA/AMPA ratios (Fig. 3). Enhanced plasticity rapidly degrades over the two weeks during which cognitive impairment develops and animals undergo terminal decline to death (Fig. 3). Acute
Acknowledgements
Work was supported by the National Institutes of Health grants R56AG052936 and R00AG034989 to GDK and a Civitan International Emerging Scholar Grant to HTV.
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