Nitric oxide (NO) has long been implicated in the generation of long-term potentiation (LTP) and other types of synaptic plasticity, a role for which the intimate coupling between NMDA receptors (NMDARs) and the neuronal isoform of NO synthase (nNOS) is likely to be instrumental in many instances. While several types of synaptic plasticity depend on NMDARs, others do not, an example of which is LTP triggered by opening of L-type voltage-gated Ca2+ channels (L-VGCCs) in postsynaptic neurons. In CA3-CA1 synapses in the hippocampus, NMDAR-dependent LTP (LTPNMDAR) appears to be primarily expressed postsynaptically whereas L-VGCC-dependent LTP (LTPL−VGCC), which often coexists with LTPNMDAR, appears mainly to reflect enhanced presynaptic transmitter release. Since NO is an excellent candidate as a retrograde messenger mediating post-to-presynaptic signaling, we sought to determine if NO functions in LTPL−VGCC in mouse CA3-CA1 synapses. When elicited by a burst type of stimulation with NMDARs and the associated NO release blocked, LTPL−VGCC was curtailed by inhibition of NO synthase or of the NO-receptor guanylyl cyclase to the same extent as occurred with inhibition of L-VGCCs. Unlike LTPNMDAR at these synapses, LTPL−VGCC was unaffected in mice lacking endothelial NO synthase, implying that the major source of the NO is neuronal. Transient delivery of exogenous NO paired with tetanic synaptic stimulation under conditions of NMDAR blockade resulted in a long-lasting potentiation that was sensitive to inhibition of NO-receptor guanylyl cyclase but was unaffected by inhibition of L-VGCCs. The results indicate that NO, acting through its second messenger cGMP, plays an unexpectedly important role in L-VGCC-dependent, NMDAR-independent LTP, possibly as a retrograde messenger generated in response to opening of postsynaptic L-VGCCs and/or as a signal acting postsynaptically, perhaps to facilitate changes in gene expression.

In the hippocampus, as in many other CNS areas, nitric oxide (NO) participates in synaptic plasticity, manifested as changes in pre- and/or postsynaptic function. While it is known that these changes are brought about by cGMP following activation of guanylyl cyclase-coupled NO receptors attempts to locate cGMP by immunocytochemistry in hippocampal slices in response to NO have failed to detect the cGMP elevation where expected, i.e. in the pyramidal neurones. Instead, astrocytes, unidentified varicose fibres and GABA-ergic nerve terminals are reported to be the prominent NO targets, raising the possibility that NO acts indirectly via other cells. We have re-investigated the distribution of cGMP generated in response to endogenous and exogenous NO in hippocampal slices using immunohistochemistry and new conditions designed to optimise cGMP accumulation and, hence, its detectability. The conditions included use of tissue from the developing rat hippocampus, a potent inhibitor of phosphodiesterase-2, and an allosteric enhancer of the NO-receptive guanylyl cyclase. Under these conditions, cGMP was formed in response to endogenous NO and was found in a population of pyramidal cell somata in area CA3 and subiculum as well as in structures described previously. The additional presence of exogenous NO resulted in hippocampal cGMP reaching the highest level recorded for brain tissue (1700 pmol/mg protein) and in cGMP immunolabelling throughout the pyramidal cell layer. Populations of axons and interneurones were also stained. According with these results, immunohistochemistry for the common NO receptor β1-subunit indicated widespread expression. A similar staining pattern for the α1-subunit with an antibody used previously in the hippocampus and elsewhere, however, proved to be artefactual. The results indicate that the targets of NO in the hippocampus are more varied and extensive than previous evidence had suggested and, in particular, that the pyramidal neurones participating in NO-dependent synaptic plasticity are direct NO targets.

Key pointsr The superior olivary complex (SOC) exhibits a spectrum of HCN1 and HCN2 subunit expression, which generate I H currents with fast and slow kinetics, respectively.r Neuronal nitric oxide synthase (nNOS) was broadly distributed across the SOC. r NO hyperpolarizes the half-activation voltage of HCN1-mediated currents and caused a slowing of the I H current kinetics in the respective nuclei (medial and lateral superior olives and superior paraolivary nucleus). This signalling was independent of cGMP.r NO also caused a depolarizing shift in the half-activation voltage of HCN2-mediated I H currents, increasing activation at resting potentials; this was cGMP-dependent.r Thus, NO signalling suppressed fast HCN1-mediated currents and potentiated slow HCN2-mediated currents, modulating the overall kinetics and magnitude of the endogenous I H .Abstract Hyperpolarization-activated non-specific cation-permeable channels (HCN) mediate I H currents, which are modulated by cGMP and cAMP and by nitric oxide (NO) signalling. Channel properties depend upon subunit composition (HCN1-4 and accessory subunits) as demonstrated in expression systems, but physiological relevance requires investigation in native neurons with intact intracellular signalling. Here we use the superior olivary complex (SOC), which exhibits a distinctive pattern of HCN1 and HCN2 expression, to investigate NO modulation of the respective I H currents, and compare properties in wild-type and HCN1 knockout mice. The medial nucleus of the trapezoid body (MNTB) expresses HCN2 subunits exclusively, and sends inhibitory projections to the medial and lateral superior olives (MSO, LSO) and the superior paraolivary nucleus (SPN). In contrast to the MNTB, these target nuclei possess an I H with fast kinetics, and they express HCN1 subunits. NO is generated in the SOC following synaptic activity and here we show that NO selectively suppresses HCN1, while enhancing I H mediated by HCN2 subunits. NO hyperpolarizes the half-activation of HCN1-mediated currents and slows the kinetics of native I H currents in the MSO, LSO and SPN. This modulation was independent of cGMP and absent in transgenic mice lacking HCN1. Independently, NO signalling depolarizes the half-activation of HCN2-mediated I H currents in a cGMP-dependent manner. Thus, NO selectively suppresses fast HCN1-mediated I H and facilitates a slow HCN2-mediated I H , so generating a spectrum of modulation, dependent on the local expression of HCN1 and/or HCN2. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.[