IN-VITRO STUDY OF H-SENSITIVE NEURONS IN THE VENTRAL MEDULLARY SURFACE OF NEONATE RATS()

We hypothesized that the direct stimulus of the central chemoreceptor neurons is the CO2/H+-induced change in intracellular pH (pHi). If it is true, pi-Ii responses during hypercapnic stimulation should be exhi bited in the central chemoreceptor neurons in the ventral medullary su rface (VMS) and some neurons in the CO2/H+ sensitive regions such as t he nucleus tractus solitarii of the medial dorsal medulla (MDM). To te st this hypothesis, the cultured VMS and MDM neurons (control) derived from one day-old neonate rats were labeled with H+-sensitive fluoresc ent indicator 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF), and were exposed to perfusate of various pHs. The H+-sensitive neuron s were determined by a rapid decrease in the intracellular BCECF fluor escence intensity. In almost all the MDM neurons (99.6%) and 94% of th e VMS neurons, the intracellular BCECF fluorescence intensity remained unchanged when the extracellular pH (pHo) was decreased. In contrast, in 0.4% of the MDM neurons (8/1800) and in 6% of the VMS neurons (111 /1800), the intracellular BCECF fluorescence intensity decreased when the pHo was decreased from 7.4 to 7.2. This subpopulation of MDM and V MS neurons were considered to be H+-sensitive neurons. The H+-sensitiv e neurons in the VMS showed positive immunoreactivity to glutamate (57 %, 17/30) and glutamic acid decarboxylase (23%, 7/30), but no immunore activity to choline acetyltransferase, tyrosine hydroxylase, phenyleth anolamine N-methyltransferase, somatostatin, serotonin and substance P . These results indicate that the H+-sensitive neurons are present spe cifically in the VMS, and are mainly glutamatergic and GABAergic. (C) 1997 Elsevier Science B.V.