The Journal of Neurophysiology Vol. 78 No. 2 August 1997, pp. 903-911
Copyright ©1997 The American Physiological Society

Factors That Reverse the Persistent Depolarization Produced by Deprivation of Oxygen and Glucose in Rat Hippocampal CA1 Neurons In Vitro

S. Yamamoto¹, E. Tanaka¹, Y. Shoji¹, Y. Kudo², H. Inokuchi¹, and H. Higashi¹

¹ Department of Physiology, Kurume University School of Medicine, Kurume 830; and ² Laboratory of Cellular Neurobiology, Tokyo University of Pharmacy and School of Life Science, Hachioji, Tokyo 192-03, Japan

Yamamoto, S., E. Tanaka, Y. Shoji, Y. Kudo, H. Inokuchi, and H. Higashi. Factors that reverse the persistent depolarization produced by deprivation of oxygen and glucose in rat hippocampal CA1 neurons in vitro. J. Neurophysiol. 78: 903-911, 1997. In CA1 pyramidal neurons in rat hippocampal tissue slices, superfusion with ischemia-simulating medium produced a rapid depolarization after 6 min of exposure. The membrane potential eventually reached 0 after 5 min (a persistent depolarization), even when oxygen and glucose were reintroduced. The role of various ions in the reversal of this persistent depolarization after reintroduction of oxygen and glucose was investigated. The peak of the persistent depolarization was decreased in solutions containing reduced Na⁺ or Ca²⁺ and in solutions containing Co²⁺ or Ni²⁺. In contrast, the depolarization was not affected by reduction of external K⁺ or Cl or by addition of tetrodotoxin (TTX), flunarizine, or nifedipine. These results suggest that sustained Na⁺ and Ca²⁺ influxes produce the persistent depolarization. The membrane potential recovered after reintroduction of oxygen and glucose in low Ca²⁺, low Cl, or K⁺-rich medium and in TTX- or tetraethylammonium-containing medium, but not in low Na⁺ or low K⁺ medium and in flunarizine- or nifedipine-containing medium. Either reduction in extracellular Ca²⁺ or addition of Co²⁺ was the most effective in promoting recovery from the persistent depolarization, suggesting that Ca²⁺ influx has a key role in causing the membrane dysfunction. The peak of the persistent depolarization was reduced by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), DL-2-amino-5-phosphonopentanoic acid (AP5), DL-amino-3-phosphonopropionic acid (AP3), or DL-amino-4-phosphonobutyric acid, suggesting that activation of non-N-methyl-D-aspartate (non-NMDA), NMDA, and metabotropic glutamate (Glu) receptors is involved in the generation and maintenance of the persistent depolarization. Among these Glu receptor antagonists, only CNQX or AP5 was able to reduce dose dependently the level of depolarization, suggesting that Ca²⁺ influx via both -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid/kainate type II receptors and NMDA receptors contributes to the membrane dysfunction. trans-1-aminocyclopentane-1,3-dicarboxylic acid (t-ACPD) did not affect the peak potential of the persistent depolarization, but it dose-dependently restored the membrane potential. AP3 antagonized the protective action of t-ACPD. The membrane potential also recovered after reintroduction when the slice was pretreated by 1,2-bis(2-aminophenoxy)ethane-N, N,N,N-tetraacetic acid tetraacetoxymethyl ester, ryanodol 3-(1H-pyrrole-2-carboxylate), 8-(diethylamino)octyl-3,4,5-trimethoxybenzoate hydrochloride, and procaine, suggesting that raised [Ca²⁺]_i from Ca²⁺-induced Ca²⁺ release pool contributes to the membrane dysfunction. It, therefore, is concluded that raised [Ca²⁺]_i has a dominant role in causing irreversible changes. The increase in [Ca²⁺]_i during the persistent depolarization may be the result of Ca²⁺ entry via both a leaky membrane and Glu-activated receptor channels as well as Ca²⁺ released from internal stores.

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