Proceedings of The Physiological Society

University of York (2002) J Physiol 539P, S007


Are GABAC receptors functionally active in the dorsal vagal motor nucleus of the rat medullary slice?

Susan A. Deuchars, Carol J. Milligan and Jim Deuchars

School of Biomedical Sciences, University of Leeds, The Worsley Building, Leeds LS2 9NQ, UK

Until recently, the GABAC receptor was thought to be functionally important only in the retina. However, numerous studies now suggest a role for this receptor in many areas of the CNS, e.g. superior colliculus (Schmidt et al. 2001). Our studies have focused on the function of the GABAC receptor in the medulla oblongata where it may play a role in cardiovascular control. In the medulla, rho;1, rho;2 and rho;3 transcripts have been detected using RT-PCR while rho;1 immunoreactivity is present in neurones in the dorsal vagal nucleus (DVN) and nucleus tractus solitarii (NTS), suggesting an important role for rho; subunits in these regions (Milligan et al. 2001). Here we determined in a medullary slice preparation the effects of applying drugs that have been selective for the GABAC receptor in the retina (see Bormann, 2000). Rats (21 days) were anaesthetised with Sagatal (60 mg kg-1) and perfused with ice-cold sucrose-ACSF. The brainstem was removed, mounted in agar and cut at 250 µm. Slices were superfused at 5 ml min-1 in ACSF bubbled with 95 % O2 and 5 % CO2 at room temperature. Whole-cell patch-clamp recordings were made from DVN neurones using patch solution with Lucifer Yellow in the pipette for recovery of recorded neurones. The effects of applications of the GABACR agonist were determined on membrane potential and input resistance. The GABAAR antagonist, bicuculline, the GABACR antagonist TPMPA or sodium pentobarbitone (50 µl in 50 ml) were pre-incubated in the ACSF before reapplying agonists. To determine the pharmacology of IPSPs elicited in the DVN, the NTS was stimulated and IPSPs isolated by applying the excitatory amino acid receptor antagonists NBQX (20-40 µM) and AP-5 (30-60 µM). The effects of TPMPA and bicuculline on these IPSPs were then determined.

Bath applications of CACA (200-500 µM) elicited depolarisations of 17.7 ± 2.3 mV (mean ± S.E.M.) in DVN neurones associated with a decrease in input resistance (n = 15). These responses were decreased in amplitude (18 ± 4.5 mV to 6.2 ± 2.8 mV, n = 5) or abolished by the GABAC antagonist TPMPA (40-160 µM, n = 7) but were also completely blocked by the GABAA receptor antagonist bicuculline (20 µM, n = 7). Applications of sodium pentobarbitone before CACA reversibly increased the amplitude of the responses to CACA by an average of 72 %. Neurones that were sensitive to CACA were filled and the slices fixed and tested for rho;1 subunit immunoreactivity (see Milligan et al. 2001). Lucifer Yellow-filled neurones were found to be immunoreactive for the rho;1 subunit and located in the DVN region of the brainstem. NTS stimulation elicited IPSPs in DVN neurones after pharmacological isolation with NBQX and AP-5. TPMPA (40-80 µM) attenuated these IPSPs (8.2 ± 2 mV to 4.9 ± 1.4 mV) while after recovery from TPMPA, bicuculline (5-20 µM) abolished the IPSP. These results indicate that CACA elicits responses in DVN neurones that are sensitive to both GABAA- and the GABAC-receptor antagonists. This may be as a result of activation of GABAC subunits.

We would like to thank The Wellcome Trust and the British Heart Foundation for their financial support.

Where applicable, experiments conform with Society ethical requirements