Effects of rapamycin on gene expression, morphology, and electrophysiological properties of rat hippocampal neurons

Abstract

SummaryPurpose-We assayed the effects of rapamycin, an immunomodulatory agent known to inhibit the activity of the mammalian target of rapamycin (mTOR) cascade, on candidate gene expression and single unit firing properties in cultured rat hippocampal neurons as a strategy to define the effects of rapamycin on neuronal gene transcription and excitability.Methods-Rapamycin was added (100 nM) to cultured hippocampal neurons on days 3 and 14. Neuronal somatic size and dendritic length were assayed by immunohistochemistry and digital imaging. Radiolabeled mRNA was amplified from single hippocampal pyramidal neurons and used to probe cDNA arrays containing over 100 distinct candidate genes including cytoskeletal element, growth factor, transcription factor, neurotransmitter, and ion channel genes. In addition, the effects of rapamycin (200 nM) on spontaneous neuronal activity and voltage-dependent currents was assessed.Results-There were no effects of rapamycin on cell size or dendrite length. Rapamycin altered expression of distinct mRNAs in each gene family on days 3 and 14 in culture. Single unit recordings from neurons exposed to rapamycin exhibited no change from baseline. When spontaneous activity was increased by blocking GABA-mediated inhibition with bicuculline, a fraction of the neurons exhibited a decreased duration of spontaneous bursts and a decrease in synaptic inputs. Rapamycin did not appear to alter voltage dependent Na+ or K+ currents underlying action potentials.Conclusions-These data demonstrate that rapamycin does not produce neurotoxicity nor alter dendritic growth and complexity in vitro and does not significantly alter voltage gated sodium and potassium currents. Rapamycin does affect neuronal gene transcription in vitro. Use of rapamycin in clinical trials for patients with tuberous sclerosis complex should involve vigilance for possible effects on seizure frequency and neurocognitive function.

Impact of rapamycin on status epilepticus induced hippocampal pathology and weight gain

“…When evaluated through a battery of acute seizure tests, rapamycin exhibited minimal acute anticonvulsant properties (Chachua et al, 2012; Hartman et al, 2012; Siebel et al, 2015) and may exhibit some pro-convulsant effects (Huang et al, 2012; Macias et al, 2013). This is not surprising, since the majority of current anti-epileptic drugs directly inhibit or activate ion channels (excitatory or inhibitory, respectively) -- mechanisms not attributed to rapamycin (Daoud et al, 2007; Ruegg et al, 2007). The mTOR pathway is, however, a key regulator of synaptogenesis (Li et al, 2010), synaptic strength (Weston et al, 2014) and long-term potentiation (Stoica et al, 2011).…”

PI3K-Akt Signaling Activates mTOR-Mediated Epileptogenesis in Organotypic Hippocampal Culture Model of Post-Traumatic Epilepsy

“…Data from acute applications of rapamycin in vitro do not suggest a direct anticonvulsant effect (Daoud et al, 2007; Rüegg et al, 2007). Structural changes such as the reduction of mossy fiber sprouting after chronic mTOR inhibition suggest an antiepileptogenic effect (Buckmaster et al, 2009; L-H Zeng NR Rensing et al, 2009; Huang et al, 2010).…”

Frontier of Epilepsy Research - mTOR signaling pathway

“…By this disruption, rapamycin fails to phosphorylate S6K1 and 4E-BP1, thus inhibits protein translation. Rapamycin itself does not seem to have any immediate effect on electrophysiological (voltage-gated sodium and potassium currents) properties of neurons in vitro (Rüegg et al, 2007). However, rapamycin has been shown to modulate the protein level of some voltage-gated potassium channels (Raab-Graham et al, 2006;Tyan et al, 2010).…”