| The auditory cortex (AC) is the highest centre of central auditory system, and the layer 5 pyramidal neurons in primary auditory cortex (AI) are importent. In vivo studies indicate that learning and experience can induce frequency-specific plasticity in AI pyramidal neurons, and the cholinergic modulation plays an important role in learning-induced cortical plasticity. However, the evidence of cellular action mechanisms is lacking. Using in vitro whole-cell patch clamp recordings, we investigated the electrophysiological properties of AI layer 5 (L5) pyramidal neurons and the effects of cholinergic receptors agonist carbachol (CCh) on the electrophysiological properties of these neurons. The results showed that a combination of the activation of cholinergic receptors and intracellular positive current injection led to persistent activity (PA) in AI L5 pyramidal neurons. Here were the detailed results:1.108 pyramidal neurons were recorded from 53 mice. Based on the firing characteristics, two types of responses were identified:regular-spiking (RS, n= 49) and intrinsic-bursting (IB, n=59). There were significant differences in the activation threshold (21.4 ± 4.0 mV vs 13.6 ± 3.5 mV) and input resistance (196.9 ± 70.2 MD vs 120.5 ± 44.2 MΩ) between RS and IB neurons (independent t-test, both p< 0.01). The percentages of neurons showing rebound depolarization were 36.7%(18/49) and 54.2% (32/59) in RS and IB neurons respectively. Hyperpolarization-activated inward current was significantly higher in IB neurons than that in RS neurons (9.8 ±3.0 pA vs 8.3 ± 4.5 pA, independent t-test,p< 0.01).2. Effects of CCh on membrane properties of pyramidal neurons.82 neurons were recorded (RS, n=40; IB, n=42). Activation of cholinergic receptors led to depolarization (82.5%,33/40 in RS neurons; 85.7%,36/42 in IB neurons), hyperpolarization (7.5%,3/40 in RS neurons; 2.4%,1/42 in IB neurons) or no change (10.0%,4/40 in RS neurons; 11.9%,5/42 in IB neurons) in membrane potentials.3. The PA, which was characterized by continuous depolarization, was induced in 79.6%(86/108) pyramidal neurons through cholinergic receptors activation and positive current injection. According to the duration, PA was classified as either long-lasting or self-terminating. The after depolarizaiton and spiking response lasted for more than 30 s in long-lasting PA while the membrane potential returned to baseline potential within 30 s in self-terminating PA. The percentages of neurons showing PA were 81.6%(40/49) in RS neurons and 78.0%(46/59) in IB neurons. And there were 67.5%(27/40) long-lasting PA in RS neurons and 76.1%(35/46) in IB neurons. The latency of PA in IB neurons was smaller than that in RS neurons (independent t-test, p< 0.05). however, there were no differences were found in the firing frequency and amplitude of plateau potential between IB and RS neurons (independent t-test, both p> 0.05).4. Further pharmacological studies showed that the PA was not dependent on the fast synaptic transmissions as it was not blocked by bath application of ionotropic glutamate receptors antagonist kynurenic acid (KA) and GAB AA receptors antagonist picrotoxin. Noticeably, the firing frequency and plateau potential of the PA were significantly increased after application of synaptic blockers (paired t-test, both p< 0.01). PA was blocked by muscarinic acetylcholine receptor (mAChR) antagonist atropine. The induction of PA depended on calcium sensitive nonspecific cationic current (ICAN) as it was blocked by ICAN antagonist flufenamic acid. Ca2+influx through L-type Ca2+-channels played roles in the induction of PA as removing extracellular Ca2+ abolished the PA and L-type Ca2+-channels blocker nifedipine (NIFE) partially blocked PA, significantly reduced spiking frequency (paired t-test, p<0.01).To sum up, there were two firing characteristics (RS and IB) in layer 5 pyramidal neurons, and there were significantly differences in the electrophysiological properties between them, this may be related to neuronic morphological structures, projection areas and the different roles in information processing. Both RS and IB neurons could induce PA, and the pharmacological studies indicated that they may share the same neuromechanism in the induction of PA. PA is independent of fast synaptic transmissions, and the induction of PA depended on the activation of mAChR, ICAN and extracellular calcium influx. L-type Ca2+-channels may palyed an important role in mediating the intracellular calcium increase. This study provided insights for understanding the action mechanisms of cholinergic modulation in central auditory system. |
PDF Full Text Request