April 18, 2014. Experimental cancer drugs called PAK inhibitors can promote dendritic spines in a genetic mouse model of psychiatric disorders, reports a study published April 3 in the Proceedings of the National Academy of Sciences. Led by Akira Sawa at Johns Hopkins University, Baltimore, Maryland, the study focuses on fixing the loss of dendritic spines, the recipients of excitatory inputs from other neurons—a deficit reported in schizophrenia and also found in mice lacking normal amounts of protein encoded by disrupted in schizophrenia 1 (DISC1). The researchers developed compounds that inhibit p21-activated kinases (PAKs), and these protected or restored spines in the mice, as well as rescued deficits in sensory gating similar to those seen in schizophrenia.
The study offers up a new candidate drug development strategy for schizophrenia. PAKs catalyze changes to cell structure and morphology, including the transformations that turn a normal cell into a cancerous one. Inhibiting PAKs might be a way to stave off the loss of dendritic spines (and their resident synapses) reported in postmortem schizophrenia brain (Glantz and Lewis, 2000). This idea has recently gotten a boost from a genetic study that implicated copy number variants containing one member of the PAK family, PAK7, in schizophrenia (see SRF related news report).
But the new study came to PAKs through a different line of inquiry—DISC1, the gene disrupted in a Scottish family enriched for mental illness. In 2010, Sawa and colleagues reported that rat neurons low in DISC1 did not respond normally to the spine-building electrical activity through the N-methyl-D-aspartate (NMDA)-type of glutamate channels: Instead of showing a spurt of spine growth, the dendrites had only sparsely scattered spines (see SRF related news report). The researchers determined that this was because kalirin-7 (Kal-7), normally tethered to DISC1, was freed in DISC1 knockdown neurons to spur spine loss via Kal-7’s interaction with Rac1, a GTPase that in turn activates PAKs. The researchers reasoned that blocking the action of PAKs could block the program of excessive pruning.
Protect and rescue
To watch the dynamic changes unfold, first author Akiko Hayashi-Takagi and colleagues confirmed their previous finding with time-lapse photography. Upon NMDA receptor activation of in vitro rat cortical neurons, spines swelled in control neurons, but shrank away in neurons transfected with DISC1 interfering RNA (RNAi).
Treatment with one of three PAK inhibitors developed by the team, however, staved off the shrinkage. When the PAK inhibitors were added to the dish an hour prior to NMDA receptor activation, no significant differences emerged in spine size between control neurons and neurons with DISC1 knockdown 20 minutes later. Two of three PAK inhibitors also prevented the loss in spine density in the DISC1 knockdown neurons.
The PAK inhibitors could also protect neurons from the ill effects of a prolonged DISC1 knockdown. Though DISC1 loss initially leads to more spines, this trend reverses with time, leading to sparser and smaller spines. Adding the PAK inhibitors at the same time as the RNAi for DISC1 resulted in spine density and size no different from neurons with control RNAi seven days later. This effect was also dose dependent and did not seem to alter normal spines in controls.
The researchers also found some evidence that the PAK inhibitors could resuscitate shrunken spines: Adding PAK inhibitors after five days of DISC1 knockdown—when the spines are presumably withered—brought about an increase in spine size, but not density, three days later.
The researchers then tested their PAK inhibitors in living mice. First, they orchestrated a DISC1 knockdown in cortical neurons of the prefrontal cortex by injecting DISC1 RNAi in utero. At postnatal day 35—mouse "adolescence"—the researchers imaged dendritic spines through a small hole in the skull by using two-photon microscopy to get a baseline. Mice with DISC1 knockdown showed about half the spine density that controls did.
However, daily injections of one of the PAK inhibitors, called FRAX486, for 25 days erased this difference. In a behavioral test of prepulse inhibition—in which the startle response to a sound is attenuated by a preceding sound—the DISC1 knockdown mice showed a strong startle—a deficit also observed in schizophrenia. But the DISC1 knockdown mice treated with FRAX486 showed less of a deficit, though not at control levels.
That PAK inhibitors could sculpt neurons and behavior alike is encouraging and provides some hope to a field running short on therapeutic ideas. But whether PAK inhibitors are viable treatments for people with schizophrenia awaits much additional research.—Michele Solis.
Hayashi-Takagi A, Araki Y, Nakamura M, Vollrath B, Duron SG, Yan Z, Kasai H, Huganir RL, Campbell DA, Sawa A. PAKs inhibitors ameliorate schizophrenia-associated dendritic spine deterioration in vitro and in vivo during late adolescence. Proc Natl Acad Sci U S A. 2014 Apr 3. Abstract