Pinpointing autism’s faulty circuitry

People diagnosed as residing on the autism spectrum suffer from a fuzzy kind of disorder. It can’t be identified by a brain scan. Not so long ago it wasn’t even thought to have a physiological basis; it was blamed on bad parenting.

Those who fit the diagnosis range from highly intelligent and articulate to non-verbal and intellectually disabled. A third suffer from seizures. They are four times more likely to have gastrointestinal problems than their peers. They may have movement difficulties, or be unusually sensitive to noise.

What unites all people with autism spectrum disorder (ASD) is a problem with their sociability, but what exactly in the brain has gone awry?People diagnosed as residing on the autism spectrum suffer from a fuzzy kind of disorder. It can’t be identified by a brain scan. Not so long ago it wasn’t even thought to have a physiological basis; it was blamed on bad parenting.

Those who fit the diagnosis range from highly intelligent and articulate to non-verbal and intellectually disabled. A third suffer from seizures. They are four times more likely to have gastrointestinal problems than their peers. They may have movement difficulties, or be unusually sensitive to noise.

What unites all people with autism spectrum disorder (ASD) is a problem with their sociability, but what exactly in the brain has gone awry?

Now, by following a hunch and using genetic tools to finely dissect the circuity of the brain, a team led by Matthew Anderson at Harvard Medical School has traced what appears to be a sociability circuit in mice.

Surprisingly the circuit does not lie in the prefrontal cortex, the part of the brain associated with communication. Rather, it is in a tiny zone at the top of the brain’s most primitive region, the brain stem. Known as the ventral tegmental area (VTA), it is well-known for its role in rewarding behaviours such as eating, having sex and taking drugs. Now it appears it may reward social behaviour as well.

Using a molecular chemogenetic switch in mice, Anderson’s team was able to turn this circuit off and on, generating animals that were asocial or hyper-social.

The findings, reported in Nature in March, may finally help open the door to new treatments for autism. “Finding a circuit that is linked to ASD is essential for developing therapeutic strategies,” says Joseph Dougherty, who studies mouse models of autism at Washington University in St. Louis.

ASD is surrounded by mysteries. The number of cases has risen dramatically. In the US its prevalence has increased from an estimated one in 2,500 children in the 1980s to one in 68 today. Other countries have shown similar increases.

There is no doubt much of that is due to the shifting way autism has been diagnosed. It only made its debut in the Diagnostic and Statistical Manual for Mental Disorders, the DSM, in 1980. One consequence is that children once categorised as intellectually disabled are now captured under the umbrella of ASD. Nevertheless many researchers suspect rates may also be rising for other reasons: children born to older parents are at greater risk, for instance. Another unsolved mystery is why five times as many boys than girls are diagnosed.

Diagnosing a child is hardly an accurate science. “We’ve lumped a range of social difficulties into the pillars of diagnosis,” says Bruce Tonge, a child psychiatrist at Monash University in Melbourne.

The latest DSM-5 relies on two pillars. The first involves a child’s ability to communicate and interact. Does he make appropriate use of language? Does she make eye contact? The second captures behaviours that are repetitive, ritualistic or show extreme sensitivities. Perhaps a child flaps his hands up and down, or is extremely distressed by the derailment of a toy train, or can’t cope with loud sounds.

A psychiatrist or multidisciplinary team examines the child usually around the age of two. Boxes are ticked in each pillar to see if there are enough to hold up a diagnosis of ASD.

Until recently genetics was not all that much help. Even now there is a clear genetic cause in only about 5% of cases. For instance, there are families where a single gene called neuroligin-3 passes on the condition. In other cases, ASD has been linked to abnormal or extra chunks of DNA on chromosome 15.

The vast majority of autism cases don’t have this clear link. Some suspect it might arise from defects in multiple genes acting together. Over the past two decades researchers have become masterful at fishing them out. One fruitful approach has been to read the DNA sequence of unaffected parents and their autistic child, to see where the child’s DNA differs. This has revealed that often the child has acquired new DNA mutations; in about 10% of cases sizable chunks of their DNA has been either duplicated or deleted – so-called ‘copy number variations’.

While hundreds of genes have been linked to ASD, they haven’t shed much light on the nature of the disorder. The genes tend to participate in global processes: for example, the CHD8 gene regulates how DNA is packaged, while the SCN2A gene codes for a sodium channel that regulates electrical transmission between brain neurons.

In his Harvard lab, faced with so many potential suspects, Anderson decided to follow a hunch about chromosome 15.  He was compelled by evidence of the yin-yang effect that followed from having too few or too many bits of this chromosome. Associated with about 2% of ASD cases, it is one of the few genetic causes of autism that can be detected under the microscope. A band on chromosome 15 is slightly bigger than it ought to be. That’s because this region, known as 15q11-13, has been triplicated. (Curiously, it is only associated with ASD when inherited from the mother.)

On the other hand, children who have lost this same bit of chromosome are the opposite of autistic; they have Angelman’s Syndrome, evidenced by a lower IQ and hyper-social behaviour. They smile and laugh, and make strong eye contact.

One of the genes that stood out in the 15q region was ubiquitin E3-ligase (Ube3a). Other trawls through the DNA of people affected by ASD were also unearthing copy number increases in the same gene. But its known functions – tagging some proteins for destruction, and turning genes on and off – provided few hints as to why it might cause autism.

So the Anderson team created mice with multiple copies of the Ube3a gene. Sure enough, the animals showed symptoms that placed them on the mouse autism spectrum. Males were less interested in new females, vocalised less, and groomed obsessively. The more copies of the gene, the more severe these behaviours became.

Deleting Ube3a had the opposite effect. Mice became more social, spending an average of 15 minutes socialising with newcomers instead of the usual five.

How was Ube3a doing this? The researchers took a look at which genes were affected by the presence of the extra copies. Genes provide recipes for proteins. It seemed Ube3a was acting like the master chef and modifying the menu. The production of about 200 dishes was being dialled down, while 400 others were ramped up.

Dauntingly complex? Not for this team of data detectives. Anderson and his colleagues checked the ASD gene databases to see which proteins on the modified menu also belonged to autism networks. They then checked to see which of the autism- and Ube3a-related proteins bound to each other. Like stepping stones, the idea was to connect the proteins in a pathway that might lead to autism.

One of the proteins dialled down is called cerebellin 1. The more copies of the Ube3a gene, the lower its production. The protein-binding network showed cerebellin 1 snapped together with two interesting autism-related genes: neurexin 1 and glutamate receptor delta subunit 1 (GRID 1). All three act at the synapse, the space between neurons where messages are relayed by neurotransmitters.

The trio form a sandwich across the synapse, with neurexin 1 at the sending side, and GRID 1 at the receiving side and cerebellin 1 in the middle. These proteins help solder particular neurons together to form a circuit.

Now the question became: what types of neurons use this triple solder? There are dozens of types, identified by the transmitters they use. To find out, the team used genetic engineering to insert extra copies of the master chef gene into specific neurons. It was only when excess Ube3a was introduced into glutamatergic or dopaminergic neurons – both associated with reward-dependent behaviours – that mice experienced social problems.

There is one tiny part of the brain, just the size of a pinhead, that has recently been shown to employ just glutamatergic and dopaminergic neurons in its circuitry: the VTA. Like tiny wires, the neurons connect to other brain areas associated with reward behaviours, such as the nucleus accumbens, the prefontal cortex and amygdala.

The hypothesis was that Ube3a was making mice asocial by taking cerebellin 1 off the menu. To find out which of the two neuron circuits was key, the researchers eliminated cerebellin 1 from the glutamatergic and then the dopaminergic neurons. When glutamatergic neurons were deprived of cerebellin, that crippled the sociability of mice.

Interestingly, the scientists found strong seizures also knocked down cerebellin 1 levels, making mice asocial. Anderson suspects seizures trigger some forms of autism, particularly Dravet’s syndrome.

Through fine sleuthing work, the team had traced sociability to a single circuit in a pinhead-sized part of the brain. But the researchers didn’t leave it at that. To prove the circuit regulated social behaviour, they developed a way to turn it on or off in mice using chemogenetics. They engineered a ‘switch’ into the glutamatergic neurons of the VTA – a receptor that responded to a chemical called clozapine-N-oxide (CNO). In the off position, the mice remained socially impaired. When fed CNO, they behaved more socially.

Localising the autism defect in these mice to the fine circuitry of the VTA is “a beautiful piece of work”, Dougherty says.

So could a treatment for ASD be a matter of toggling the sociability switch?  Anderson believes it might. “It has a therapeutic flavour,” he says. “Someday we might be able to translate this into a treatment that will help patients.”