碩士 / 國立臺灣大學 / 藥理學研究所 / 96 / Orexins are a novel family of two hypothalamic peptides, consisting of orexin A and orexin B, which were also named hypocretin 1 and hypocretin 2, respectively. They were identified to be the endogenous agonists of an orphan GPCR in 1998, which was, therefore, named orexin receptor and consists of OX1 and OX2 receptors. OX1 receptors display higher affinity for orexin A than orexin B, while OX2 receptors have similar affinity for both orexin A and orexin B. Orexin-containing neurons are mostly localized in the lateral hypothalamus and send projections widely through the brain, including the hippocampus. Orexin A had been reported to affect learning and memory tasks and hippocampal synaptic plasticity in vivo, mostly in the rat, as well as affected synaptic plasticity of hippocampal slices of rats and mice in vitro. However, controversial results were reported. We, therefore, investigated the effects of orexins on the synaptic plasticity, long-term potentiation (LTP) and depotentiation, at Schaffer Collateral-CA1 synapses of mouse hippocampal slices. Field excitatory postsynaptic potentials (fEPSPs) were evoked by stimulating the Schaffer collateral pathway and recorded in the stratum radiatum of the CA1 region of hippocampal slices. LTP was induced by high frequency stimulation (HFS) which consisted of 3 trains of theta burst stimulation (TBS), separated by 15s. Each TBS consisted of 4 pulses at 100 Hz, separated by 200 ms. Depotentiation was induced by low frequency stimulation (LFS, 1Hz, 15 min) 40 min after HFS.
Orexin A (30-300 nM) had no effect on the baseline fEPSPs after 10 min treatment. Orexin A treated 10 min before and during HFS, decreased HFS-induced LTP dose-dependently. The magnitude of LTP (% of the baseline fEPSPs) 40 min after HFS was reduced by pretreatment with orexin A 100 nM and 300 nM. The magnitude of PTP (% of the baseline fEPSPs) 40 min after HFS was mild dose-dependently reduced by pretreatment with orexin A 30 nM, 100 nM and 300 nM. Depotentiation induced by LFS (expressed by the % restoration of the magnitude of LTP) 40 min after LFS was not different with orexin A.
[Ala11, D-Leu15]-orexin B (3~1000nM) also had no effect on the baseline fEPSPs after 10 min treatment. [Ala11, D-Leu15]-orexin B treated 10 min before and during HFS, decreased HFS-induced LTP dose-dependently. The magnitude of LTP (% of the baseline fEPSPs) 40 min after HFS was reduced by pretreatment with [Ala11, D-Leu15]-orexin B 30 nM, 300 nM and 1000 nM. The magnitude of PTP (% of the baseline fEPSPs) 40 min after HFS had no different by pretreatment with [Ala11, D-Leu15]-orexin B. Depotentiation induced by LFS (expressed by the % restoration of the magnitude of LTP) 40 min after LFS was not different with [Ala11, D-Leu15]-orexin B.
SB 334867 (3 μM), a selective OX1 receptor antagonist, partially antagonized the effect of orexin A. The magnitude of LTP (% of the baseline fEPSPs) in slices treated with orexin A+ SB-334867 was higher in slices with 100 nM orexin A, which was significantly lower than the control slices. The magnitude of PTP (% of the baseline fEPSPs) in slices treated with 100 nM orexin A had no significantly difference in slices with orexin A+ SB-334867, which was not significantly different with the control slices. SB-334867 alone had no effect on basal transmission, PTP, LTP and depotentiation.
SB 334867 (3 μM) slightly antagonized the effect of [Ala11, D-Leu15]-orexin B. The magnitude of LTP (% of the baseline fEPSPs) in slices treated with 300 nM [Ala11, D-Leu15]-orexin B + SB 334867 was higher in slices with [Ala11, D-Leu15]-orexin B, which was significantly lower than the control slices. The magnitude of PTP (% of the baseline fEPSPs) in slices treated with 300 nM [Ala11, D-Leu15]-orexin B had no statistically significant difference in slices with [Ala11, D-Leu15]-orexin B + SB 334867, which was not significantly different with the control slices. SB 334867 alone had no effect on basal transmission, PTP, LTP and depotentiation.
Compound 29 (30 μM), a selective OX2 receptor antagonist, partially antagonized the effect of 100 nM orexin A. The magnitude of LTP (% of the baseline fEPSPs) in slices treated with 100 nM orexin A + Compound 29 was higher in slices with 100 nM orexin A, which was significantly lower than the control slices. The magnitude of PTP (% of the baseline fEPSPs) in slices treated with 100 nM orexin A was no different in slices with orexin A + Compound 29, which was no significantly difference with the control slices. Compound 29 alone had no effect on basal transmission, PTP, LTP and depotentiation.
Compound 29 (30 μM) slightly antagonized the effect of [Ala11, D-Leu15]-orexin B. The magnitude of LTP (% of the baseline fEPSPs) in slices treated with [Ala11, D-Leu15]-orexin B + Compound 29 (133±7.1%, n=5) was higher in slices with 300 nM [Ala11, D-Leu15]-orexin B (120.8±7.2%, n=5), which was significantly lower than the control slices (141.9±3.5%, n=16). The magnitude of PTP (% of the baseline fEPSPs) in slices treated with 300 nM [Ala11, D-Leu15]-orexin B (146.8±8.7%, n=5) was not significantly different in slices with [Ala11, D-Leu15]-orexin B + Compound 29 (186.5±28%, n=5), which had no significantly difference with the control slices (181.5±10.4 %, n=16). Compound 29 alone had no effect on basal transmission, PTP, LTP and depotentiation.
Paired-pulse facilitation (PPF) of fEPSPs, a presynaptic phenomenon resulting from Ca2+ accumulation in the presynaptic terminals in response to stimulation by a pair pulses with short interval is induced by paired-pulse (50 ms interval) stimulation. The PPR during LTP decreased, as compared to basal PPF ratio (PPR), suggesting that increased presynaptic transmission plays a role in the LTP evoked by TBS in CA1 region of mouse hippocampal slices although this LTP is mostly mediated by NMDA receptors. Orexin A (100 nM) did not affect basal PPR but slightly reversed the PPR decreased during LTP.
It is concluded that orexin A and [Ala11, D-Leu15]-orexin B decreased LTP but not affect depotentiation in CA1 region of mouse hippocampal slices. The impairment of LTP is mediated by both OX1 and OX2 receptors. The efficacy mediated by OX1 receptors is higher than that by OX2 receptors. It is suggested that presynaptic inhibition plays a little role in orexin-induced impairment; instead, postsynaptic NMDA receptor inhibition might contribute to this impairment through PLC/PKC pathway activated by OX1 and OX2 receptor activation.
| Identifer | oai:union.ndltd.org:TW/096NTU05550008 |
| Date | January 2008 |
| Creators | Chia-Hsu Lee, 李家旭 |
| Contributors | 邱麗珠 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 62 |
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