D NO and Ryanodine Receptors

It has been reported that ryanodine receptors are present in axons (Ouardouz et al., 2003), which raises the interesting possibility that NO might affect axonal physiology via this route. There are so far no studies of the effects of NO on axonal ryanodine receptors, but guidance might be taken from studies in muscle. Here, NO is believed to play a role in the normal physiological function of muscle cells

(i.e., contraction), where ryanodine receptors play a key role (Stamler and Meissner, 2001; Salama et al., 2000; Hare, 2003). In muscle, ryanodine receptors are well-established components in the calcium-induced calcium release mechanism, whereby calcium ions entering a muscle cell via its voltage-dependent, L-type calcium channels triggers the release of calcium ions from the sarcoplasmic reticulum (SR). The release is achieved by opening the high conductance SR calcium release channel, the ryanodine receptor. These receptors are exceptionally rich in free thiol groups, and certain of these have a high sensitivity to oxidation/ nitrosation, resulting in channel opening (Fig. 5) (Xu et al., 1998; Stoyanovsky et al., 1997; Eu et al., 1999). Indeed, S-nitrosation of up to 12 sites on the receptor can lead to the progressive activation of the channel, which is reversed by denitrosation (Xu et al., 1998), although nitrosation of a single cysteine may occur physiologically (Eu et al., 2000). In the initial studies, rather high, seemingly nonphysiological concentrations of exogenous nitric oxide were used to study these effects, but it is now emerging that within the normal

Figure 5 Records showing the effects of NO (in the form of the donor GSNO) on the cardiac ryanodine receptor located in a planar lipid bilayer. Single-channel currents are shown as upward deflections from the closed levels, before (top record) and 1 minute after 1 mM GSNO (middle record) and then 1 minute after 10 mM dithiothreitol (DTT) (bottom record). Exposure to NO increased the frequency of channel opening, which was reversed by DTT. (Modified from Xu et al., 1998.)

muscle cell NO formation can occur in distinct microdomains where nNOS (NOS1) and eNOS (NOS3) iso-forms occur in close proximity with ryanodine receptors and SR calcium ATPase, and with L-type calcium channels, respectively (reviewed in Hare, 2003). In this way, NO is delivered as required in the immediate vicinity of its intended targets.

In muscle cells, NO can nitrosate critical thiols on the ryanodine receptor to effect activation and/or inhibition of the receptor, and these actions appear to play a role in normal muscle contraction. Indeed, NO cycles in the beating heart on millisecond time scales, and in canine heart cells, the ryanodine receptor is reported to be endogenously nitrosated (Xu et al., 1998; Eu et al., 2000). By analogy with the observations in muscle, it seems reasonable to propose on the basis of the observations by Ouardouz et al. (2003) that NO may promote calcium release in axons from intra-axonal stores. This may be especially prominent in inflammatory MS lesions, with sustained high concentrations of NO and axons unadapted for such NO exposure. When considered in conjunction with the fact that calcium concentrations within neuronal endoplasmic reticulum can be substantial, especially after electrical activity (Pozzo-Miller et al., 1999), it is clear that axons could become flooded with damaging concentrations of calcium. Indeed, whereas muscle cells contain high concentrations of the NO-scavenger myoglobin, this is not the case in axons, which may therefore be more sensitive to NO.

Apart from affecting the function of ryanodine receptors by the direct nitrosation of critical thiols, there is also evidence that NO may affect their function by phosphorylation via cGMP-dependent protein kinase (Suko et al., 1993). Also, apart from calcium release via ryanodine receptors, it is possible that inositol-1,4,5-triphosphate (IP3) receptors could be involved, although the effects of NO on IP3 receptors have not been extensively studied.

VIII. NO and the Na+/K+ ATPase (Sodium Pump)

Na+/K+ ATPase plays a key role in axonal physiology, being responsible for sodium extrusion after impulse activity, and contributing to the maintenance of resting potential. Indeed, Na+/K+ ATPase consumes approximately 50% of the energy supply in the CNS. It is significant, therefore, that NO is quite potent in impairing the function of the enzyme, both in vitro (Guzman et al., 1995; Sato et al., 1995; Muriel and Sandoval, 2000) and in vivo (Liu and Sheu, 1997) when NO production is raised by iNOS formation. Indeed, there is now evidence that the Na+/K+ ATPase is nitrosated under physiological conditions in vivo as a result of nNOS activity, even without the additional contribution from iNOS (Jaffrey et al., 2001). The inhibition appears to be due to an interaction

Figure 5 Records showing the effects of NO (in the form of the donor GSNO) on the cardiac ryanodine receptor located in a planar lipid bilayer. Single-channel currents are shown as upward deflections from the closed levels, before (top record) and 1 minute after 1 mM GSNO (middle record) and then 1 minute after 10 mM dithiothreitol (DTT) (bottom record). Exposure to NO increased the frequency of channel opening, which was reversed by DTT. (Modified from Xu et al., 1998.)

between NO or a related reactive nitrogen species and a thiol group at the active site of the enzyme (Sato et al., 1997; Boldyrev et al., 1997), although other evidence also supports an action mediated via cGMP (McKee et al., 1994). Brain Na+/K+ ATPase is more vulnerable to inhibition than that from kidney (Boldyrev et al., 1997). If, as seems likely, function of the Na+/K+ ATPase is impaired in axons affected by inflammatory lesions in MS, the axons may become depolarized, and also prone to sodium loading, thereby becoming predisposed to degeneration (see later).

0 0

Post a comment