Abstract:
GABA (4-aminobutyrate) is synthesized through the decarboxylation of LGlu-
(L-Glu-+ H+ ---> GABA + C02), and compared to many free amino acids is
present in high concentrations in plant cells. GABA levels rise rapidly and
dramatically in response to varied stress conditions including anaerobiosis.
Recent papers suggest that GABA production and associated H+ consumption are
parts of a metabolic pH-stat mechanism which ameliorates the intracellular pH
decline associated with anaerobiosis or other treatments. To test this hypothesis
GABA production and efflux have been measured in isolated Asparagus sprengeri
cells in response to three treatments which potentially cause intracellular
acidification. Acid loads were imposed using 60 min of (i) anaerobiosis, (ii) H+/LGlu-
cotransport, and (iii) treatment with permeant weak acids (butyric, acetic and
propionic). Both intra- and extracellular GABA concentrations increased more than
100% after anaerobiosis, almost 1000% after H+/L-Glu- cotransport (light or dark)
and almost 5000/0 after addition of 5 mM butyric acid at pH 5.0. HPLC analysis of
amino acids indicates that as GABA concentrations increased in response to
butyric acid addition, glutamate concentrations decreased. Time-course studies
demonstrated that added butyric acid stimulates GABA production by 2800/0 within
15 seconds. A fluorescent determination of cytosolic pH indicates that addition of
butyric or other weak acids resulted in a rapid reduction in cytosolic pH of 0.6 pH
units. The half time for the response to butyric acid addition is 2.1 seconds,
indicating that the decline in cytosolic pH is rapid enough to account for the rapid
stimulation of GABA production. The acid load in response to butyric acid addition
was assayed by measurements of 14C-butyric acid uptake. Calculations indicate
that GABA production accounted for 45% of the imposed acid load. The biological
significance of GABA efflux is not yet understood. The results support the original
hypothesis suggesting a role for GABA production in cellular pH regulation.
