Acetylcholine raises excitability by inhibiting the fast transient potassium current in cultured hippocampal neurons.

Abstract

The effects of acetylchoLine on cultured hippocampal neurons were investigated by using the whole-cell version of the patch-clamp technique. The CAl region of the hippocampus was excised from brain slices ofyoung rats (12-19 day old), incubated in a papain solution, and dissociated. Neurons were plated on a glial feeder layer. The experiments were conducted mostly on neurons cultured for 2-6 days. Upon depolarization under voltage clamp, these cells exhibited a fast transient outward current (A-current), which was inhibited by 4-aminopyridine (2.5 mM). Acetylcholine (0.1 p) also inhibited this A-current, as did the muscarinic agonists bethanechol and muscarine. As expected from their inhibition of the A-current, acetylcholine and 4-aminopyridine both increased the amplitude of the action potential and prolonged its duration. We conclude that the inhibition of the A-current constitutes a mechanism by which acetylcholine exerts its excitatory influence on hippocampal neurons.Neuronal excitation produced by the muscarinic action of acetylcholine (AcCho) is an important cellular event for brain function, but details of its ionic mechanism are not fully understood. In 1971 Krnjevid et al. (1) investigated the muscarinic action of AcCho on cortical neurons by using intracellular microelectrodes, and they concluded that AcCho probably produces its excitatory action by reducing both resting and delayed potassium currents. Recently, Brown and Adams (2) and Halliwell and Adams (3) analyzed the muscarinic action under voltage clamp and explained muscarnnic excitation as being due primarily to an inhibition of a potassium current, which they named the M-current.We have investigated the action of cholinergic agonists on cultured hippocampal neurons by using the whole-cell patchclamp technique. Our results show that cholinergic agonists inhibit the fast transient potassium current, which was designated as the A-current by Connor and Stevens (4). This inhibition of the A-current would play a role in cholinergic excitation. The effect of AcCho resembles that of 4-aminopyridine (APd), which has been shown to be a relatively specific inhibitor ofthe A-current (5). Previously, it has been shown that thyrotropin releasing hormone (6), noradrenaline (7), and the intracellular level of cyclic AMP (8, 9) may modify neural activity via their effects on A-current. MATERIALS AND METHODSCulture. The culture method was similar to that developed for dissociated cell cultures of locus coeruleus (10) and nucleus basalis (11). Young rats postnatal,(12)(13)(14)(15)(16)(17)(18)(19) day old; Charles River Breeding Laboratories) rather than newborn or embryonic rats were used in the main, because it is known that muscarinic receptors in the hippocampus begin to increase after the birth (12). Coronal sections (400 Am thick) of brains were obtained with a vibratome. From these brain slices, the CA1 region of hippocampus was excised under a dissecting microscope. We began the work by using trypsin dissociation (10, 11), but we soon realized ...

Rapid desensitization of glutamate receptors in vertebrate central neurons.

“…Neurons from either 6-day embryonic chicken spinal cord or 2-to 7-day postnatal rat hippocampus were dissociated with 0.01% trypsin or 0.1% papain, respectively, as described (8,9). They were grown on glial monolayers in Eagle Fine-tipped ionophoretic electrodes were filled with 250 mM glutamate at pH 8. Sibling electrodes, filled with 3 M KCl, measured 60-80 MQ.…”

Cholinergic modulation of synaptic transmission and plasticity in entorhinal cortex and hippocampus of the rat

“…ACh depolarizes cells, facilitates burst ®ring and induces slow excitatory postsynaptic potential (EPSP) by blocking potassium channels. 7,12,17,28,29,32 In contrast, ACh suppresses neurotransmitter release 18 and excites inhibitory neurons. 31 In addition, cholinergic activation promotes development of theta rhythm.…”

Elevation of cytosolic calcium by cholinoceptor agonists in SH‐SY5Y human neuroblastoma cells: estimation of the contribution of voltage‐dependent currents

“…Whilst SH-SY5Y cells do express L-and N-type voltage-sensitive Ca2t channels, they do not appear to be involved in carbacholstimulated Ca2t entry . On the other hand, muscarinic agonists are capable of exciting neurones, generally by inhibiting K+-currents (Nakajima et al, 1986;North, 1986) …”