Suppression of the Nicotinic Acetylcholine Response in Rat Superior Cervical Ganglionic Neurons by Steroids

Abstract : The effects of various types of steroids on the nicotinic acetylcholine (ACh) receptor (nAChR)-mediated responses were investigated in superior cervical ganglionic neurons acutely dissociated from rats using nystatin perforated patch recording. ACh induced a peak followed by a gradual decrease in the inward current at a holding potential of -40 mV. Nicotine, but not muscarine, mimicked ACh. Hydrocortisone at a concentration of > 10^-6 M reversibly suppressed both the peak and steady-state nicotine-induced currents ( I _nic) in a noncompetitive manner. The inhibition of I _nic by hydrocortisone did not show any voltage dependency and persisted in the presence of either cyclic AMP modulators, forskolin and 3-isobutyl-1-methylxanthine, or a protein kinase A inhibitor, N -[2-(methylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride (H-89). β-Estradiol, androsterone, aldosterone, and 17α-estradiol mimicked hydrocortisone in its inhibitory action on ACh-induced currents ( I _ACh). The potency for the inhibitory actions on I _Ach was as follows : androsterne > β-estradiol > hydrocortisone ≥ aldosterone =17α-estradiol. Cholesterol had no effect on the I _ACh. In conclusion, the structural characteristics of steroid are thus considered to be necessary to block nicotinic I _ACh in rat superior cervical ganglionic cells, whereas the cholesterol side chain might disturb the inhibitory action of the steroid skeleton on nAChRs.     

Proadrenomedullin N-Terminal 20 Peptide (PAMP), an Endogenous Anticholinergic Peptide: Its Exocytotic Secretion and Inhibition of Catecholamine Secretion in Adrenal Medulla

Abstract: In cultured bovine adrenal medullary cells, stimulation of nicotinic receptors by carbachol evoked the Ca²⁺-dependent exocytotic cosecretion of proadrenomedullin N-terminal 20 peptide (PAMP) (EC₅₀ = 50.1 µ M ) and catecholamines (EC₅₀ = 63.0 µ M ), with the molar ratio of PAMP/catecholamines secreted being equal to the ratio in the cells. Addition of PAMP[1–20]NH₂ inhibited carbachol-induced ²²Na⁺ influx via nicotinic receptors (IC₅₀ = 2.5 µ M ) in a noncompetitive manner and thereby reduced carbachol-induced ⁴⁵Ca²⁺ influx via voltage-dependent Ca²⁺ channels (IC₅₀ = 1.0 µ M ) and catecholamine secretion (IC₅₀ = 1.6 µ M ). It did not alter high K⁺-induced ⁴⁵Ca²⁺ influx via voltage-dependent Ca²⁺ channels or veratridine-induced ²²Na⁺ influx via voltage-dependent Na⁺ channels. PAMP seems to be a novel antinicotinic peptide cosecreted with catecholamines by a Ca²⁺-dependent exocytosis in response to nicotinic receptor stimulation.     

Autoradiographic Localization of Angiotensin II Receptor Binding Sites on Noradrenergic Neurons of the Locus Coeruleus of the Rat

The locus coeruleus (LC) of the rat was lesioned by microinjection of selective neurotoxins into the brainstem. 6-Hydroxydopamine (6-OHDA), 3 micrograms/microliter, given unilaterally at two sites 0.6 mm apart on the rostro-caudal axis of the LC, was used to lesion catecholamine-containing neuronal elements. Ibotenic acid, 2.5 micrograms/0.5 microliters, administered similarly was used to lesion nerve cell bodies. Two weeks after administration of the neurotoxin, lesion efficacy was determined based on the norepinephrine content of the cerebral cortex ipsi- and contralateral to the lesion. 6-OHDA lesions of the LC caused a 46% reduction in ipsilateral cortical norepinephrine and a 60% reduction in specific 125I-[Sar1, Ile8]-angiotensin II (125I-SIAII) binding in the LC. Ibotenic acid lesions of the LC caused a 73% reduction in ipsilateral cortical norepinephrine and a 81% reduction in specific 125I-SIAII binding in the LC. These results indicate that AII receptor binding sites in the LC are localized on noradrenergic nerve cell bodies or their dendritic and axonal ramifications within the LC.     

Properties of Acetylcholine Receptor Ion Channels in the Acutely Dissociated Neurons of the Marginal Division in the Rat Striatum

Cell-attached mode of patch clamp technique was employed to investigate the properties of acetylcholine (ACh)-induced ion channels in acutely dissociated neurons from the marginal division (MrD) of rat striatum. Two types of conductance states (25 pS and 60 pS) were recorded. The 25 pS channel (more than 80%) was the main type in the neurons of MrD and was described here. The amplitudes of inward currents increased with hyperpolorization and the reversing potential was about 0 mV. Both single short opening and long burst openings were observed in MrD neurons. Two time constants of these two kinds of ion channels are 0.29 ms, 1.84 ms and 1.96 ms, 18.24 ms, respectively. Average close time can be fitted with two exponential functions, the two time constants are 1.7 ms and 54 ms. Probability of channel opening is about 0.012 and no voltage-dependence was found. The properties of reversing potential, voltage-independence and the form of agonist to the ion channels indicated that the recorded channel currents flow through AChR channels. The mAChR is involved in slow synaptic transmission and Ach can not induce the opening of mAChR ion channel. The binding site of ACh to AChR and the nAChR ion channel are the same protein, ACh can only activate nAChR ion channel directly. Therefore, the recorded ion channels in the present study are nAChR ion channels. The results suggest that nAChR ion channels exist in the neurons of MrD and the MrD probably is involved in learning and memory mechanism of the brain.