Inhalational anesthetics are known to disrupt PDZ2 domain-mediated protein-protein interactions of the postsynaptic density (PSD)-95 protein. The aim of this study is to investigate the underlying mechanisms in response to early isoflurane exposure on synaptic PSD-95 PDZ2 domain disruption that altered spine densities and cognitive function. The authors hypothesized that activation of protein kinase-G by the components of nitric oxide signaling pathway constitutes a mechanism that prevents loss of early dendritic spines and synapse in neurons and cognitive impairment in mice in response to disruption of PDZ2 domain of the PSD-95 protein.


Postnatal day 7 mice were exposed to 1.5% isoflurane for 4 h or injected with 8mg/kg active PSD-95 wild-type PDZ2 peptide or soluble guanylyl cyclase activator YC-1 along with their respective controls. Primary neurons at 7 days in vitro were exposed to isoflurane or PSD-95 wild- type PDZ2 peptide for 4 h. Co-immunoprecipitation, spine density, synapses, cyclic guanosine monophosphate dependent protein kinase activity, and novel object recognition memory were assessed.


Exposure of isoflurane or PSD-95 wild-type PDZ2 peptide relative to controls causes: (1) decrease in PSD-95 co-immunoprecipitate relative to N-methyl-D-aspartate receptor NR2 subunits precipitate (mean±SD[in percentage of control]): isoflurane (54.73±16.52), P=0.001 and PSD-95 wild-type PDZ2 peptide (51.32±12.93), P=0.001; (2) loss in spine-density (mean±SD [spine density per 10 micrometer]): control (5.28 ± 0.56 ) versus isoflurane (2.23±0.67), P<0.0001, and PSD-95 mutant PDZ2 peptide (4.74±0.94) versus PSD-95 wild-type PDZ2 peptide (1.47±0.87), P<0.001; and decrease in synaptic puncta (mean±SD [in percentage of control]): isoflurane (41.1±14.38), P=0.001 and PSD-95 wild-type PDZ2 peptide (50.49±14.31), P<0.001. NO donor or cyclic guanosine monophosphate analogue prevents the spines and synapse loss and decline in the cyclic guanosine monophosphate-dependent protein kinase activity but this prevention was blocked by soluble guanylyl cyclase or protein kinase-G inhibitors in primary neurons; (3) deficits in object recognition at 5 weeks: (mean±SD[recognition index]): male; control (64.08±10.57) versus isoflurane (48.49±13.41), P=0.001, (n=60) and female; control (67.13±11.17) versus isoflurane (53.76±6.64), P=0.003 (n=58). Isoflurane-induced impairment in recognition memory was preventable by the introduction of YC-1.


Activation of soluble guanylyl cyclase or protein kinase-G prevents isoflurane or PSD-95 wild-type PDZ2 peptide induced loss of dendritic spines and synapse. Prevention of recognition memory with YC-1, a NO independent activator of guanylyl cyclase supports a role for the soluble guanylyl cyclase mediated protein kinase-G signaling in countering the effects of isoflurane-induced cognitive impairment.