Oxygensensitive K Channels in PC 12 Cells

The outward Ko2 current that is selectively and reversibly inhibited by hypoxia in PC 12 cells is carried by a Kv channel. Ko2 currents have been characterized in other chemosensitive cells. In most chemosensitive cells such as carotid body type I cells and rat pulmonary artery smooth muscle cells , the Ko2 current is voltage-dependent; however, in rat carotid body and in neuroepithelial body, a background K+ current was also recently proposed to constitute an 02-sensitive K+ conductance (1,3, 11, 14). At least four types of K+ channels are expressed in PC 12 cells, distinguished by their unitary current conductances and current-voltage relationships: a small conductance K+ channel (Ksm;14 pS), a Ca2+-activated K+ (KCa) channel (102 pS) and twoK+ channels with similar conductance (20 pS; Fig. 2A-B) (7, 15). These last two channels differ in their time-dependent inactivation: one is a slowly-inactivating channel (Kdr), while the other belongs to the family of fast transient K+ channels (Ktr). Patch clamp studies revealed that hypoxia selectively inhibits only the 20 pS slowly-inactivating delayed-rectifier type of K+channel in PC 12 cells (7). Figure 2C shows the single-channel properties of the 20 pS Ko2 channel recorded in a cell-attached patch in PC12 cells and the effect of hypoxia on the channel activity. Single-channels currents were elicited by step-pulse depolarization in normoxia and 2 min after hypoxia. The inhibitory effect ofhypoxia was apparent in the decreased amplitude of the ensemble-averaged currents due to a decrease in open probability (P0) of the channels, with no change in conductance (Fig. 2C, bottom panels) (7). The effect ofhypoxia on the Ko2 channel in PC 12 cells as well as CB type I cells and other recombinant Ko2 channels is manifested by a similar mechanism involving a decrease in channel P0, a slowing of activation kinetics, and little effect on channel closing (7, 11, 23). Interestingly, the sensitivity to hypoxia of the channel is maintained in inside-out patch configuration both in PC12 cells and CB type I cells, suggesting that cytosolic soluble factors might not be required for the hypoxic response (7, 11).

Figure 2. Characterization of single Ko2 channels in PC 12 cells. A: The unitary current-voltage (i/V) relationships of a delayed-rectifier (Kdr) channel and a transient (Ktr) Kv channel which have similar conductance but different time-dependent properties. The i/V relationships were obtained using a 800 ms ramp pulse depolarization from a holding potential (HP) of -60 mV to +50 mV (2.8 mM K+ in the pipette). The straight lines indicate the slope conductance for Kdr (19 pS) and Ktr (20 pS). Outward K+ currents (ensemble-average currents) of Kdr and Ktr channels, elicited by a 180 ms step pulse from -60 to +50 mV in the same patch, respectively, are shown in right panels. B: The i/V relationships of the small conductance (Ksm) Kv channel and a large conductance Kq, channel show that the slope conductances are 15 and 102 pS for K$m and Kq, channels, respectively. C: Hypoxia inhibits the activity of the 20-pS Kdr channel. Top panels show representative currents, elicited by 180 ms step depolarizing pulses from -60 to +50 mV before (normoxia) and after 2-min exposure to hypoxia (10% 02). The corresponding ensemble-averaged currents (from 100 consecutive traces) are shown in bottom panels. Dashed lines represent the zero current. The slope conductance, measured by ramp depolarization, is shown

Figure 2. Characterization of single Ko2 channels in PC 12 cells. A: The unitary current-voltage (i/V) relationships of a delayed-rectifier (Kdr) channel and a transient (Ktr) Kv channel which have similar conductance but different time-dependent properties. The i/V relationships were obtained using a 800 ms ramp pulse depolarization from a holding potential (HP) of -60 mV to +50 mV (2.8 mM K+ in the pipette). The straight lines indicate the slope conductance for Kdr (19 pS) and Ktr (20 pS). Outward K+ currents (ensemble-average currents) of Kdr and Ktr channels, elicited by a 180 ms step pulse from -60 to +50 mV in the same patch, respectively, are shown in right panels. B: The i/V relationships of the small conductance (Ksm) Kv channel and a large conductance Kq, channel show that the slope conductances are 15 and 102 pS for K$m and Kq, channels, respectively. C: Hypoxia inhibits the activity of the 20-pS Kdr channel. Top panels show representative currents, elicited by 180 ms step depolarizing pulses from -60 to +50 mV before (normoxia) and after 2-min exposure to hypoxia (10% 02). The corresponding ensemble-averaged currents (from 100 consecutive traces) are shown in bottom panels. Dashed lines represent the zero current. The slope conductance, measured by ramp depolarization, is shown

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