Hippocampal neuronal maturation triggers post-synaptic clustering of brain temperature-sensor TRPV4 Koji Shibasaki a, * , Makoto Tominaga b, c, d , Yasuki Ishizaki a a Department of Molecular and Cellular Neurobiology, Gunma University Graduate School of Medicine, Maebashi 371-8511, Japan b Division of Cell Signaling, National Institute for Physiological Sciences, Okazaki 444-8787, Japan c Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, Okazaki 444-8787, Japan d Department of Physiological Sciences, The Graduate University for Advanced Studies, Okazaki 444-8585, Japan article info Article history: Received 5 January 2015 Available online 28 January 2015 Keywords: TRPV4 Hippocampus Development Synapse Cerebellum PSD-95 abstract Compartmentalization of neuronal function is achieved via specifically localized clustering of ion channels in discrete subcellular membrane domains. Transient receptor potential (TRP) channels exhibit highly variable cellular and subcellular patterns of expression. We previously revealed that the thermo- sensor TRPV4 (activated above 34  C) is gated by physiological brain temperatures in hippocampal neurons and thereby controls their excitability. Here, we examined synaptic clustering of TRPV4 in developing hippocampal neurons. We found that TRPV4 accumulated in the soma of immature hippo- campal neurons, and did not localize to post-synaptic locations although PSD-95-labeled post-synaptic structures were evident. During the maturation of neurons, TRPV4 was targeted to dendrites and also clustered at post-synaptic locations. Taken together, we reveal that TRPV4 localizes to post-synaptic sites and the post-synaptic targeting is strictly regulated in a neuronal maturation-dependent manner. © 2015 Elsevier Inc. All rights reserved. 1. Introduction The hippocampus contains neural circuitry that is crucial for higher brain functions, such as learning and memory [1]. The neu- rons that form circuits in the hippocampus require transmembrane cation influx for depolarization and action potential generation [2,3]. Many kinds of ion channels, such as voltage-gated Na þ channels and Ca 2þ channels, contribute to membrane depolarization [2,4]. TRPV4 is a nonselective cation channel, first described as an osmosensor detecting hypotonic stimuli, that shares 40% amino acid identity with TRPV1 [5e8]. TRPV4 can also be activated by heat (i.e., temperatures >27e34  C), the phorbol ester derivative 4a-PDD (4a- phorbol 12,13 didecanoate), and lipid metabolites [9e12]. TRPV4 was reported to be necessary for the response to changes in osmotic pressure and functions as an osmotic sensor in the CNS [13,14]. We found that TRPV4 was strongly expressed in hippocampal neurons and constitutively activated by physiological brain temperature to control neuronal excitability [15]. Furthermore, we reported that TRPV4 is also expressed in microglia and specific subtypes of astrocytes in the brain, where it regulates synaptic activity through gliotransmitter release [16,17]. These results strongly indicate that TRPV4 is a key molecule regulating neuronal excitability. It is well known that small changes in brain temperature affect brain function [18e21]. Clinical evidence suggests that brain tem- perature is particularly a critical determinant of cognitive function. For example, it has been reported that therapeutic hypothermia leads to cognitive dysfunction in heart disease patients [22], while cooling of the prefrontal cortex causes an impairment in the per- formance of a delayed matching-to-sample task [23]. In rodents, brain cooling significantly impairs spatial learning [24]. These re- ports indicate that the maintenance of brain temperature is important for healthy brain function. We have already revealed an involvement of the thermo-sensor function of TRPV4 in brain function following its activation by brain temperature changes [15e17]. In this study, we examined the synaptic targeting of TRPV4 in developing hippocampal neurons. 2. Materials and methods 2.1. Animals TRPV4-deficient (TRPV4KO) mice were generated by crossing heterozygous mice, and the genotypes were determined by PCR as * Corresponding author. Department of Molecular and Cellular Neurology, Gunma University Graduate School of Medicine, Maebashi 371-8511, Japan. Fax: þ81 27 220 7955. E-mail address: shibasaki@gunma-u.ac.jp (K. Shibasaki). Contents lists available at ScienceDirect Biochemical and Biophysical Research Communications journal homepage: www.elsevier.com/locate/ybbrc http://dx.doi.org/10.1016/j.bbrc.2015.01.087 0006-291X/© 2015 Elsevier Inc. All rights reserved. Biochemical and Biophysical Research Communications 458 (2015) 168e173