Ionic storm in hypoxic/ischemic stress: can opioid receptors subside it?

Abstract

Neurons in the mammalian central nervous system are extremely vulnerable to oxygen deprivation and blood supply insufficiency. Indeed, hypoxic/ischemic stress triggers multiple pathophysiological changes in the brain, forming the basis of hypoxic/ischemic encephalopathy. One of the initial and crucial events induced by hypoxia/ischemia is the disruption of ionic homeostasis characterized by enhanced K(+) efflux and Na(+)-, Ca(2+)- and Cl(-)-influx, which causes neuronal injury or even death. Recent data from our laboratory and those of others have shown that activation of opioid receptors, particularly delta-opioid receptors (DOR), is neuroprotective against hypoxic/ischemic insult. This protective mechanism may be one of the key factors that determine neuronal survival under hypoxic/ischemic condition. An important aspect of the DOR-mediated neuroprotection is its action against hypoxic/ischemic disruption of ionic homeostasis. Specially, DOR signal inhibits Na(+) influx through the membrane and reduces the increase in intracellular Ca(2+), thus decreasing the excessive leakage of intracellular K(+). Such protection is dependent on a PKC-dependent and PKA-independent signaling pathway. Furthermore, our novel exploration shows that DOR attenuates hypoxic/ischemic disruption of ionic homeostasis through the inhibitory regulation of Na(+) channels. In this review, we will first update current information regarding the process and features of hypoxic/ischemic disruption of ionic homeostasis and then discuss the opioid-mediated regulation of ionic homeostasis, especially in hypoxic/ischemic condition, and the underlying mechanisms.

Keywords: Ca2+ channels; K+ channels; Na+ channels; hypoxia/ischemia; ionic homeostasis; neuroprotection; opioids; δ-opioid receptor.

Figure 1. Anoxia induced changes in extracellular potassium concentration ([K⁺]e ) and extracellular DC potential (ΔDC) in mouse cortical slice

Note that anoxia largely increased the concentration of extracellular potassium (upper trace) and decreased DC potential (lower trace). The changes in K⁺ were distinctly characterized by a two-phase response to hypoxia/ischemia. The first phase shows a relatively slow increase in [K⁺]e starting after 20–60 sec of hypoxia/ischemia. Over several minutes of anoxia, [K⁺]e increase reached a threshold level (no more than the so-called “K⁺ ceiling” level), phase 2 was characterized by a rapid, abrupt and large increase in [K⁺]e, accompanied by a negative shift in extracellular DC potential. Within about 20 seconds of the onset of phase 2, [K⁺]e reached a peak value, then gradually decreased. Upon return to normoxia, [K⁺]e fell rapidly, and often undershot to below the basal levels before steadily returning to the pre-anoxia level.

Figure 2. Effect of DOR activation by UFP 512 on anoxic disruption of K⁺ homeostasis

Trace recordings of A: Control (Cont), B: UFP 512 (1 µM) (UFP 512-1.0), and C: UFP 512 (10 µM) (UFP 512-10). D–F are statistical results of each recording parameter. The following parameters were derived to assess K⁺ homeostasis: (1) the latency of anoxia-induced [K⁺]e increase (Latency), which was defined as a time period from the beginning of anoxia to the time point when anoxia induced a K⁺ electrode voltage change greater than 1mV; (2) maximal [K⁺]e ([K⁺]max), which was the peak change in extracellular K⁺ concentration induced by anoxia; (3) the undershooting of [K⁺]e (Undershoot), which referred to the minimal value of [K⁺]e during reoxygenation. **p<0.01, ***p<0.001 as compared with the control; ##p<0.01 in comparison to UFP 512-1.0. Note that UFP 512 at 1 µM (UFP 512-1.0) significantly attenuated anoxic increase in [K⁺]max, with a significantly prolonged latency to anoxia and a significant delay of peak increase in [K⁺]e (n=27). Increasing concentration of UFP 512 up to 10 µM (UFP 512-10) did not further attenuate the anoxia-induced disruption of K⁺ homeostasis except for the response latency that was prolonged (n=15).

Figure 3. DOR, but not MOR, activation is protective against anoxia-induced disruption of K⁺ homeostasis in the cortex

Trace recordings of A: Control, B: DADLE (1.0 µM) (DADLE1.0), C: DADLE (10 µM) (DADLE10), D: UFP 512 (1 µM), and E: DAGO (10 µM). Note that anoxia-induced increase in [K⁺]max was significantly attenuated by DOR activation with two naturally distinct DOR agonists, DADLE and UFP 512. Whereas MOR activation by DAGO did not produce any significant change in anoxia-induced increase in [K⁺]max. These results, therefore, confirm DOR, but not MOR, activation is protective of cortical neurons against anoxia-induced disruption of K⁺ homeostasis.

Figure 4. Paradigm showing mechanisms of DOR attenuation of anoxic/ischemic disruption of K⁺ homeostasis

Our data showed that the absence of extracellular Ca²⁺ and blockade of Ca²⁺-activated K⁺ channels (BK) greatly attenuated the anoxia-induced disruption of K⁺ homeostasis, suggesting that the attenuation of DOR activation against anoxic disruption of K⁺ homeostasis in the cortex is likely attributed to an inhibition of hypoxia-induced increase in Ca²⁺ entry-BK channel activities. Besides, DOR-induced protection against the anoxic K⁺ responses was largely abolished by low Na⁺ perfusion with either impermeable N-methyl-D-glucamine or permeable Li⁺, substitution, blockade of voltage-gated Na⁺ channels and NMDA receptor channels. Whereas Non-NMDA receptor channels and Na⁺/Ca²⁺ exchangers, though involved in anoxic disruption of K⁺ homeostasis in certain degrees, are less likely the targets of DOR signals. PKC-dependent, but PKA-independent, pathway is also involved in the attenuation of anoxic disruption of K⁺ homeostasis in the cortex because we could not demonstrate the involvement of PKA in the DOR protection against anoxic increase in extracellular K⁺ because blocking PKA with H89 did not result in any changes in the DOR effect. In sharp contrast, PKC inhibition with chelerythrine reversed, and PKC activation by PMA mimicked, the protective effect of DOR activation.