Australian Black Rock scorpion toxin may help solve mystery of chronic pain


Researchers at UC San Francisco and the University of Queensland have discovered a scorpion toxin that targets the “wasabi receptor,” a chemical-sensing protein found in nerve cells that’s responsible for the sinus-jolting sting of wasabi and the flood of tears associated with chopping onions.

Because the toxin triggers a pain response through a previously unknown mechanism, scientists think it can be used as a tool for studying chronic pain and inflammation, and may eventually lead to the development of new kinds of non-opioid pain relievers.

The scientists isolated the toxin, a short protein (or peptide) that they dubbed the “wasabi receptor toxin” (WaTx), from the venom of the Australian Black Rock scorpion.

The discovery came as the researchers were conducting a systematic search for compounds in animal venom that could activate, and therefore be used to probe and study, the wasabi receptor—a sensory protein officially named TRPA1 (pronounced “trip A1”) that’s embedded in sensory nerve endings throughout the body.

When activated, TRPA1 opens to reveal a channel that allows sodium and calcium ions to flow into the cell, which can induce pain and inflammation.

“Think of TRPA1 as the body’s ‘fire alarm’ for chemical irritants in the environment,” said John Lin King, a doctoral student in UCSF’s Neuroscience Graduate Program and lead author of a study published August 22, 2019 in Cell, which describes the toxin and its surprising mode of action. “When this receptor encounters a potentially harmful compound—specifically, a class of chemicals known as ‘reactive electrophiles,’ which can cause significant damage to cells—it is activated to let you know you’re being exposed to something dangerous that you need to remove yourself from.”

Cigarette smoke and environmental pollutants, for example, are rich in reactive electrophiles which can trigger TRPA1 in the cells that line the surface of the body’s airway, which can induce coughing fits and sustained airway inflammation.

The receptor can also be activated by chemicals in pungent foods like wasabi, onions, mustard, ginger and garlic—compounds that, according to Lin King, may have evolved to discourage animals from eating these plants. WaTx appears to have evolved for the same reason.

Though many animals use venom to paralyze or kill their prey, WaTx seems to serve a purely defensive purpose. Virtually all animals, from worms to humans, have some form of TRPA1.

But the researchers found that WaTx can only activate the version found in mammals, which aren’t on the menu for Black Rock scorpions, suggesting that the toxin is mainly used to ward off mammalian predators.

“Our results provide a beautiful and striking example of convergent evolution, whereby distantly related life forms – plants and animals – have developed defensive strategies that target the same mammalian receptor through completely distinct strategies,” said David Julius, Ph.D., professor and chair of UCSF’s Department of Physiology, and senior author of the new study.

But what the researchers found most interesting about WaTx was its mode of action. Though it triggers TRPA1, just as the compounds found in pungent plants do – and even targets the very same site on that receptor – the way it activates the receptor was novel and unexpected.

First, WaTx forces its way into the cell, circumventing the standard routes that place strict limits on what’s allowed in and out. Most compounds, from tiny ions to large molecules, are either ingested by the cell through a complex process known as “endocytosis,” or they gain entry by passing through one of the many protein channels that stud the cell’s surface and act as gatekeepers.

But WaTx contains an unusual sequence of amino acids that allows it to simply penetrate the cell’s membrane and pass right through to the cell’s interior.

Few other proteins are capable of the same feat.

The most famous example is an HIV protein called Tat, but surprisingly, WaTx contains no sequences similar to those found in Tat or in any other protein that can pass through the cell’s membrane.

“It was surprising to find a toxin that can pass directly through membranes.

This is unusual for peptide toxins,” Lin King said. “But it’s also exciting because if you understand how these peptides get across the membrane, you might be able to use them to carry things – drugs, for example – into the cell that can’t normally get across membranes.”

Once inside the cell, WaTx attaches itself to a site on TRPA1 known as the “allosteric nexus,” the very same site targeted by pungent plant compounds and environmental irritants like smoke.

But that’s where the similarities end.

Plant and environmental irritants alter the chemistry of the allosteric nexus, which causes the TRPA1 channel to rapidly flutter open and closed.

This allows positively charged sodium and calcium ions to flow into the cell, triggering pain. Though both ions are able to enter when TRPA1 is activated by these irritants, the channel exhibits a strong preference for calcium and lets much more of it into the cell, which leads to inflammation.

By contrast, WaTx wedges itself into the allosteric nexus and props the channel open. This abolishes its preference for calcium. As a result, overall ion levels are high enough to trigger a pain response, but calcium levels remain too low to initiate inflammation.

To demonstrate this, the researchers injected either mustard oil, a plant irritant known to activate the wasabi receptor, or WaTx into the paws of mice.

With mustard oil, they observed acute pain, hypersensitivity to temperature and touch – key hallmarks of chronic pain – and inflammation, as evidenced by significant swelling. But with WaTx, they observed acute pain and pain hypersensitivities, but no swelling.

“When triggered by calcium, nerve cells can release pro-inflammatory signals that tell the immune system that something’s wrong and needs to be repaired,” Lin King said.

“This ‘neurogenic inflammation’ is one of the key processes that becomes dysregulated in chronic pain. Our results suggest that you can decouple the protective acute pain response from the inflammation that establishes chronic pain.

Achieving this goal, if only in principle, has been a longstanding aim in the field.”

The researchers believe their findings will lead to a better understanding of acute pain, as well as the link between chronic pain and inflammation, which were previously thought to be experimentally indistinguishable.

The findings may even lay the groundwork for the development of new pain drugs.

“The discovery of this toxin provides scientists with a new tool that can be used to probe the molecular mechanisms of pain, in particular, to selectively probe the processes that lead to pain hypersensitivity,” Lin King said. “And for those interested in drug discovery, our findings underscore the promise of TRPA1 as a target for new classes of non-opioid analgesics to treat chronic pain.”

Pain is a physiologically important phenomenon as it alerts an organism to tissue damage or potential tissue damage [1]. Pain is initiated when peripheral terminals of a subgroup of sensory neurons, termed nociceptors, are activated to produce action potentials [2]. This depolarization of nociceptors is produced by specialized pain receptors that detect various chemical, thermal, and mechanical noxious stimuli [2,3].

The pain signal is then transmitted to the spinal cord dorsal horn and eventually to higher regions in the central nervous system (CNS) where it is processed [4]. Subsequently, an appropriate response to the noxious stimulus is generated to avoid further injury [1,5].

Moreover, the memory of pain deters the affected organism from repeating actions that evoke this unpleasant experience [6].However, this pain sensation following exposure to noxious stimuli (i.e., acute pain) could be undesirable when undergoing a medical procedure or when the pain is too intense and debilitating following injury [7].

Chronic pain is another instance in which suppression of the nociceptive system is required. Chronic pain is defined as a sensation of pain that persists long after the expected healing of the underlying injury when pain is no longer serving any useful role [8,9,10].

Indeed, chronic pain is among the leading causes of seeking medical attention, accountable for about 20% of patients in primary care [11]. This pain pathology can stem from nerve damage (neuropathic pain) or be associated with conditions that produce continuous stimulation of the pain pathway, such as inflammation [8,9,12].

Chronic pain can be accompanied by plastic changes to nerves leading to altered detection, transmission, processing, and regulation of pain [13,14]. These impairments generate an abnormal and hyperexcitable function of the nociceptive system, leading to persistent and intensified pain sensations [13].Currently, treatment of chronic pain is lacking as the available drugs achieve only partial analgesia and in just a fraction of the patients [14].

To date, most analgesics in use target ion channels and receptors in the spinal cord and brain.

Thus, these agents modulate the transmission and processing of the pain signal centrally [15].

Additionally, the targets of these drugs are involved in processes other than nociception [16].

For example, opiates provide varying degrees of efficacy in the treatment of different pain types by activating opioid receptors in spinal and supra-spinal domains [13].

Due to their central activity, these agents are notorious in producing serious adverse effects, including respiratory depression, sedation, euphoria, dependence, and addiction [13,16].

While opiates also produce peripheral unwanted effects, these CNS-related side effects are especially concerning as opioid abuse and opioid-related deaths have gained epidemic proportions in the United States. Thus, pain pathologies in which opiates are also moderately effective (e.g., neuropathic pain) are preferably treated with atypical analgesics (e.g., pregabalin, duloxetine, amitriptyline) [14].

However, most of these analgesics commonly in use were initially developed and are prescribed for the treatment of other diseases such as depression and epilepsy while also presenting poor selectivity to their targets in some cases [14,15,17].

Due to this, patients experience numerous side effects when treated with these agents [16].

These adverse effects may reduce the compliance to the pharmacological therapy and further contribute to the failure of pain management. Thus, there is a dire need for novel, safe, and efficacious analgesics for the treatment of chronic pain.Receptors and ion channels in the peripheral terminals and axons of nociceptors were shown to be pivotal in the generation of pain [1,2,18]. It is possible that more effective analgesia could be achieved by targeting transduction and transmission in nociceptors, thus blocking pain at its source [19].

Additionally, analgesics with a peripheral site of action can exert an improved safety profile.

This can be achieved by targeting proteins that are expressed selectively in nociceptors [16]. A

nother avenue is to design agents that cannot penetrate the blood-brain barrier into the CNS. Indeed, there is a growing effort in the search for new analgesics that act peripherally [20]. Transient receptor potential vanilloid 1 (TRPV1) and transient receptor potential ankyrin 1 (TRPA1) are two pain receptors that emerged as potential targets for such analgesics [20].

These cation channels are activated by numerous noxious stimuli from many sources including inflammatory mediators and were suggested to have a role in the detection of noxious temperature [19]. As pain pathologies often involve altered sensitivity to heat or cold, suppressing TRPV1 and TRPA1 activation could be a promising approach [20]. Acid-sensing ion channels (ASICs) are another pain receptors that are drawing attention in this context as these cation channels (with high preference to sodium) were shown to be involved in inflammatory pain and chronic pain conditions [21,22]. Blocking the action potential propagation through the nociceptor axon by modulating voltage-gated sodium channels (NaV) could also be highly effective in alleviating pain [23]. It was found that several NaV channels are important in evoking action potentials in nociceptors where they are selectively expressed [24].

Thus, specific attenuation of the pain signal could be obtained by inhibiting these channels.The development of new modulators requires deep understanding of both the structure and function of ion channels.

Natural toxins can be used to gain such insights as they affect functionally essential domains in ion channels [6,25]. Additionally, toxins have evolved to be stable, potent and specific to proteins that are physiologically significant [6,26].

These features highlight the importance of toxins in identifying new targets for pharmacological intervention and in the design of novel drugs.

Moreover, toxins can be used as lead compounds in the process of drug development or as drugs themselves [21,27]. A striking example for this is ziconotide, a synthetic version of a toxin found in the venom of the cone snail Conus magus, which was approved by the FDA in 2004 for the treatment of severe refractory chronic pain [13].

Due to the peptidic nature of this toxin, it has to be injected intrathecally where it inhibits the N-type voltage-gated calcium channels through binding to their α1Bsubunit [13,28,29].

By inhibiting these pre-synaptic channels in the central terminals of nociceptors, ziconotide reduces the release of pro-nociceptive neurotransmitters thereby disrupting the transmission of pain signals in the spinal cord [13].

As evoking an aversive response could be a useful tool in the defensive arsenal of venomous organisms, the pharmacopeia libraries that are venoms contain numerous toxins known to modulate nociceptive targets and probably many more such toxins that are yet to be identified. Undeniably, these toxins were and will be instrumental in understanding the nociceptive system.In this review, we will focus on plant and animal toxins targeting the aforementioned prominent ion channels that are peripherally expressed in nociceptors (Figure 1).

We will evaluate the contribution of these toxins to the study of the structure and function of these channels. Additionally, toxins’ potential role in the design of novel ion channel modulators aiming at analgesia will be discussed.

Toxins 11 00131 g001 550

Figure 1. Schematic representation of plant and animal toxins targeting ion channels involved in pain. The following represents only a partial list of toxins that have been found to modulate the activity of TRPV1, TRPA1, ASIC, and NaV channels.

More information:Cell (2019).

Journal information: Cell
Provided by University of California, San Francisco


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