**4. Discussion**

doses of THC containing products are associated with an increased risk of developing respiratory infections [119]. However, it has been difficult to establish a clear relationship between smoked cannabis and more severe lung disorders, such as cancer, since tobacco use is often co-morbid with cannabis. While in general the number of smoked cannabis cigarettes is lower on a per day basis, this risk cannot be overlooked. Additionally, the inhalation delivery methods for cannabis smoke are somewhat more diverse than for tobacco and include pipes, water pipes, burning on metal and vaporizers. All of these delivery methods produce smoke with its own set of chemical characteristics that depend on the temperature at which the smoke was created and any filtering that occurred before the smoke was inhaled. Processing of the plant material into oils or resins before combustion adds another level of complexity to the potential

Zebrafish larvae are an established model for testing vertebrate toxicity, teratogenicity and environmental risk assessment [120–122]. The use of these models has proven to be a valuable resource for testing the toxicity of various extracts and condensates obtained from tobacco cigarette smoke [6, 123, 124]. It was found that smoke from tobacco cigarettes was more toxic and produced different phenotypes than that of nicotine alone, suggesting, that the other toxic components found in tobacco smoke are having an effect. The use of the previously validated smoke testing models for testing cannabis smoke has the potential to provide information on

One of the major complexities of working with cannabis is the fact that it is currently known to be comprised of 500+ constituents and more than 100 cannabinoid molecules [125]. The potential of interactions between many of these compounds is high and has been widely demonstrated for THC and CBD. It has been shown that the zebrafish can be used to assess the interaction of THC and CBD with respect to their effects on locomotor activity along with the uptake and metabolism of each compound [106]. Many of the aforementioned zebrafish models have the potential to be used to assess the potential interaction of THC and CBD and likely other cannabinoids. This is important as one of the next steps in the use of the zebrafish models for testing cannabinoids is to begin to test various extracts and isolates from cannabis for their bioactivity. Having an understanding of how the pure compounds interact in an *in vivo* system and how this relates to their activity in complex mixtures derived from plant material will be extremely valuable. In addition to the interaction of the various compounds found within cannabis, the use of cannabis or cannabinoids as therapeutics is also complicated by the fact that many patients are already taking a prescription drug for their particular indication. The zebrafish testing platforms appear

Understanding the pharmacokinetics and pharmacodynamics of cannabis is complicated by the fact that CBD (and potentially other cannabinoids) has numerous targets and mechanisms of action that contribute to its various biological effects. Similar to CBD a high percentage of neuroactive compounds have multiple targets and act on them within similar concentration ranges. This polypharmacology has both advantages and disadvantages. As many disease etiologies are not entirely known and may be multifactorial, there may be a substantial benefit of having activity on multiple targets. However, this may also increase the side effect potential and the potential

chemical diversity of the smoke that is inhaled.

22 Recent Advances in Cannabinoid Research

developmental, cardiac, behavioral/neural and acute toxicity.

to have the potential to characterize some of these interactions as well.

**3.7. Multi-drug interactions and polypharmacology**

One of the unique characteristics of researching the effects of cannabis and cannabinoids using various models and for various disease indications is that often there is already clinical data on the effects in humans. While much of the data is often anecdotal in nature, it does allow for animal model testing to be used to back validate the findings of the clinical trials. By designing top-down translational research studies we can begin to elucidate the biological basis of the clinical findings and potentially provide information on the mechanism of action of therapeutic compounds. This is particularly true for cannabis uses where the cannabinoid mechanism of action is often difficult to discern. The use of animal models of disease may help to elucidate these mechanisms and further define the etiology of the disease.

As outlined in this chapter, zebrafish have an established endocannabinoid system that is highly analogous to that of humans. Additionally, as a model system both adults and larval zebrafish provide numerous models of disease that have been shown to be efficacious for testing the therapeutic potential of novel compounds. Importantly the response patterns in these various disease models following exposure to different cannabinoids reveal unique characteristics for each cannabinoid. Thus far, only a limited number of these models have been used to test the efficacy of cannabinoids. However, the framework is in place for an expansion of the use of zebrafish in this field.
