Adenosine A1 Receptors Are Involved In The Modulation Of The Rhythmical Respiration And The Discharge Activities Of Respiratory Neurons In Neonatal Rat Brainstem Slice In Vitro

Object: 1) This experiment was expected to explore the effect of adenosine A1-receptors on the central respiratory rhythm.; 2) To test whether adenosine A1 receptors exerted a significant effect on the discharge activities of biphasic expiratory neurons and inspiratory neuronsMethods: Experiments were performed in in vitro brainstem slice preparations from neonatal rats (0~2d). These preparations include the medial region of the nucleus retrofacialis (mNRF); a part of pre-Botinger Complex, ventral respiratory group (VRG) and dorsal respiratory group (DRG). 1) Respiratory rhythmical discharge activity (RRDA) were recorded from hypoglossal nerve rootlets before and during perfusion of the slice preparation with adenosine A1 receptor agonist R-phenylisopropyl-adenosine (R-PIA) and its specific antagonist 8-cyclopentyl-1,3- dipropylxanthine (DPCPX) into a modified Kreb's perfusion solution (MKS); 2) As monitoring the respiratory-related burst activities of hypoglossal nerve rootlets, we recorded the discharge of biphasic expiratory neurons /inspiratory neurons in the mNRF by inserting the glass microeletrode into the mNRF of the slice. These biphasic expiratory neurons were characterized by the discharge activity restricted in the late expiratory phase and the early expiratory phase of the following cycle as well as inhibition in the inspiratory phase. The inspiratory neurons was identified by their on-going activities that were correlated with RRDA of the hypoglossal nerve. Drugs were administered by bath applicationand their effects on the neuronal activities were investigated.Results: 1. The discharges of hypoglossal nerve rootlets were recorded: Application of 0.1 % DMSO had no effect on the discharge activities. Control experiments were performed without drug application. Bath application of 2 umol/L DPCPX for 10 min decreased the respiratory cycle (RC) significantly by 27.07% (P<0.05) and shortened the expiratory time (TE) by 28.38% (P<0.05), whereas no significant change on the inspiratory time (TI) or discharge integral amplitude (IA) were observed. These effects were partially reversible after washout of DPCPX . Bath application of R-PIA 10 umol/L for 20 min prolonged RC by 58.49% (P<0.01) and extended TE by 63.14% (P<0.01), but shortened TI by 27.18% (P<0.01) and decreased IA by 30.22% (PO.01). 2) After application of R-PIA for 15 min, RC and TE were significantly prolonged (P<0.01) with obvious decreases in TI and IA (P<0.01), compared with the control. These effects were partially reversed by additional administration of DPCPX.2. The rhythmical discharges of respiratory neurons and activities of the hypoglossal nerve rootlets were simultaneously recorded: 1) From the biphasic expiratory neurons : Application of DPCPX significantly shortened the burst intervals of hypoglossal nerve by 29.81%, altered the discharge activity of biphasic expiratory neurons with a 28.27% decrease in the expiratory time (TE) and a 20.12% increase in the amplitude of integrated (IA) (p<0.05). Concurrently, the peak discharge frequency (PFn) was increased by 48.13% and the discharge frequency (Fn) in the whole expiratory phase also significantly increased. After washout of DPCPX, the respiratory activities were recovered to the control level. By comparision with DPCPX, R-PIA produced opposite effects on the hypoglossal nerve and the biphasic expiratory neurons. Moreover, there was no significant difference between the effects of R-PIA on the respiratory discharges without prior application of DPCPX and following perfusion of DPCPX and washout; 2) From the inspiratory neurons: Similar to the effects on the biphasic expiratory neurons, DPCPX also produced a decrease in RP (31.10%) and TE (32.52%) of inspiratory neurons (p<0.05) without any significant changes in inspiratory time (TI, 2.35%) and IA (2.69%). Application of R-PIA increased RP by 38.81%, shortened TI by 34.12 (p<0.05) and also decreased the PFn of inspiratory neurons by 37.75% (p<0.05). However, the effect ofDPCPX and R-PIA on Fn was significant in the middle 300 ms phase of inspiration, but not in the initial and terminal 100 ms phases.Contusion: The results indicated that in neonatal rat brainstem slice in vivo: 1) Adenosine A1-receptors are involved in the modulation of rhythmical respiration in mammals; 2) Adenosine Al receptors are involved in the phase-switching between expiration and inspiration by affecting biphasic expiratory neurons; 3) Adenosine Al receptors may modulate the inhibitory synaptic inputs from inspiratory neurons.