Excitatory synapses onto PV-INs are capable of undergoing plasticity30-32
, and this study provides the first molecular mechanism for a postsynaptic form of plasticity at these synapses. Activity-dependent expression of Narp by presynaptic excitatory neurons regulates homeostatic adaptations of circuit activity by enhancing the strength of excitatory synapses on PV-INs, concomitantly increasing their network-driven firing rate. PV-INs are the most abundant subtype of interneuron within the hippocampus19
and are implicated in processes such as gamma oscillations33
, visual cortical plasticity34
, and fear memory resilience35
. Additionally, the dysfunction and/or loss of PV-INs may underlie several neurological disorders such as temporal lobe epilepsy36
. Therefore, uncovering the molecular mechanisms of how these neurons regulate the strength of their synapses has implications for understanding plasticity and cognitive disorders.
The present model of Narp-dependent synaptic plasticity is consistent with its regulation as an IEG, and its ability to bind AMPARs and sugars7, 15
. Increases in activity increase Narp expression in excitatory neurons and Narp is subsequently secreted, and preferentially accumulates, at excitatory synapses on PV-INs. Narp is required for activity-regulated changes in GluR4-mediated synaptic strength at these synapses. By this process, an IEG can evoke transsynaptic effects to modulate circuit activity. It is interesting to contrast the respective roles of Narp
in homeostatic scaling. With sustained increases in activity, Arc is rapidly expressed and functions to promote the endocytosis and downregulation of GluR1 in pyramidal neurons4, 5
. At the same time, Narp targets to PV-INs where it functions to cluster GluR4 and potentiate excitatory synapses on these neurons. Using independent mechanisms, Narp
thereby function in a complementary manner, at two separate populations of glutamatergic synapses, to reset pyramidal neuron activity back to baseline levels.
Despite these molecular insights, several important questions arise. First, how is Narp
expression regulated by activity? Recent evidence demonstrates that the activity-dependent transcription of Narp
mRNA requires Ca2+
influx through L-type voltage-gated calcium channels (VGCCs) and subsequent activation of Ca2+
/calmodulin (CaM), CaM-dependent kinases, and Extracellular signal-regulated kinase (ERK) 1/238
. Additionally, Narp
is misregulated by knockdown of the neuronal IEG transcription factor Npas4 via RNAi expression39
. It is interesting to note that Npas4
, and Narp
share many similar features: both are activity-regulated genes, require Ca2+
influx through L-type VGCCs for their induction, are expressed primarily in excitatory neurons, and yet are both key regulators of the inhibitory network39
. Whether Npas4 is required for Narp expression remains to be studied.
Second, how does Narp selectively accumulate on PV-INs? Glycoproteins appear to be important for the targeting of Narp to excitatory synapses on PV-INs, consistent with the general property of pentraxins to bind sugars40
. It is notable, however, that perineuronal nets are not localized precisely at synapses41
. One model consistent with current data envisions that secreted Narp accumulates locally within the glycoprotein network, aided by lectin-based interactions. Narp might then diffuse along pre or postsynaptic membranes and localize to the synapse by interactions with other sugars or proteins, or form disulfide linked complexes with NPR11
. The lectin properties of pentraxins are linked to the Ca2+
binding pocket, which in turn, is important for proper folding of the pentraxin domain40
. Accordingly, it is difficult to selectively disrupt lectin properties by mutagenesis. Simple addition of recombinant Narp to cultures does not result in selective binding to PV-INs (unpublished observation), suggesting that targeting may involve processes beyond simple lectin-dependent binding.
Important questions also arise regarding how Narp can evoke an increase of GluR4, but not other AMPARs, on PV-INs. Binding of Narp to AMPARs does not require sugar adducts to the receptor, but rather, appears to be dependent on protein sequences within the N-terminal X-domain10
. Previous reports demonstrate the capacity for NPs to bind and cluster AMPARs at sites of cell-cell contact8, 9, 14
. It is possible that Narp-containing pentraxin complexes preferentially retain GluR4 on the cell surface, and this conjecture is consistent with the observation that NPs cluster homomeric AMPARs consisting of GluR4 subunits better than any other AMPAR subunit9
. However, the avidity of Narp binding for GluR1 and GluR4 is not dramatically different in binding assays (unpublished observation) suggesting that the difference in activity-dependent accumulation may be due to factors in addition to their association with Narp, such as the level of GluR expression in PV-INs or other selective protein interactions. Currently, assays of native GluR4 trafficking are technically limited due to lack of appropriate antibodies, and our attempts at expressing an N-terminal-tagged GluR4 transgene resulted in similar elevated levels in both WT and Narp−/−
PV-INs which were unresponsive to activity.
The observation that Narp−/−
mice showed accelerated kindling to class V seizures provides insight into conditions in which Narp may contribute to the suppression of network excitability. It is notable that the baseline properties of excitatory synapses on hippocampal PV-INs in the dentate gyrus were not different in Narp−/−
mice; only after MECS did we observe a change in excitatory input strength. The activity level in vivo
was much less than in our primary cultures and this difference was consistent with a minimal role of Narp under basal conditions in the hippocampus in vivo. Narp−/−
mice showed identical initial responses to kindling stimuli but clearly diverged in their responses as the kindling process evolved. Together, this suggests that Narp becomes part of the physiological adaptation only under conditions of intense activity, and is consistent with culture models that suggest the inability to recruit GluR4 levels to excitatory synapses on PV-INs in response to activity may underlie enhanced kindling. These studies do not exclude a role for Narp in scaling of PV-INs in physiological plasticity, since recordings of sEPSCs may not detect the small subset of Narp-associated excitatory synapses relevant for proper network function under basal conditions. Moreover, studies of Narp
mRNA expression indicate that Narp is induced by non-epileptiform activity in models of cocaine administration42
, monocular deprivation7
, and in vivo
. Because of the striking complexity of interneuron populations in vivo
and their importance to integrated neural function it will be compelling to assess the contribution of Narp-dependent homeostatic plasticity in broader studies of physiological plasticity and models of disease.