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The most widely recognized feature of the Helodermatidae family is that its members are venomous. Misunderstanding and confusion about this trait have accompanied Heloderma since before the genus was described by Hernandez in 1577. As discussed in chapter 1, the specific epithet, suspectum, was chosen for the Gila Monster because E.D. Cope suspected it was venomous (Cope 1869), but it was not until well into the twentieth century that scientists agreed that, indeed, this was true. Debate continues over whether the venom is used for prey acquisition or defense, and, more recently, promise has arisen over its potential applications in modern medicine and pharmacology.
In this chapter, I outline the structure and function of the venom delivery system of helodermatid lizards. I provide a historical overview, synthesize recent developments in venom biochemistry, and consider the function of the venom system in its ecological context. I conclude the chapter with an overview of envenomation history and the treatment of bites.
Several authors in the latter half of the nineteenth century commented on the "vile nature" of Heloderma and recognized that its unique grooved dentition might be associated with a venomous bite (see Bogert and Martin del Campo 1956). Butit was not until the 1880s that the anatomy of the venom gland was investigated (Fischer 1882), and experiments were conducted to show that its salivary secretions were, in fact, toxic to other animals (Mitchell and Reichert 1883). Instead of resolving uncertainty about the venomous nature of Heloderma, however, this work served to initiate a controversy that lasted for several decades. Subsequent experiments (Yarrow 1888) failed to show toxic effects of Heloderma venom, probably because inappropriate methods were used to collect the venom. Additional investigators examined the histology of the venom gland (Holm 1897) and showed, with controls, that carefully extracted venom injected into small vertebrates had lethal effects (Santesson 1897; Van Denburgh 1898; Van Denburgh and Wight 1900). Nevertheless, skepticism remained. Snow (1906), having suffered a bite without experiencing serious pain or swelling, again raised the question of whether Heloderma was venomous. In 1907, Goodfellow made the following statement: "... exhaustive studies were made by some of the attaches of the Smithsonian Institution, among whom was Dr. R. E. Shufeldt, concerning the nature of the animal, and conclusions reached which the writer had previously attained-that the reptile was nonvenomous; and it may be accepted as conclusively demonstrated that the bite of the "monster" is innocuous per se. In 1913, an authoritative, 244-page book summarizing detailed studies of the venom of Heloderma was published by the Carnegie Institution of Washington (Loeb et al. 1913). This book contained detailed studies by 11 contributors on numerous aspects of the venom, including its biochemistry and effects on various physiological systems in a number of organisms. By the 1920s, this book, and additional studies on the venom gland, venom effects, and venom characteristics by Phisalix (1911, 1912, 1917, 1922), finally convinced the scientific community that helodermatid lizards were indeed venomous, and the debate was settled over the "suspected" venomous nature of Heloderma.
VENOM DELIVERY SYSTEM
Heloderma delivers venom through an efficient system consisting of paired venom glands that empty through ducts at the base of venom-conducting teeth. Venom is produced and housed in multilobed venom glands (fig. 9), which, unlike those of venomous snakes, are located in the lower jaw and drain through ducts associated with each of the lobes. In contrast, the venom glands of snakes are situated behind the eye, above the upper jaw, and drain through a single duct that leads to an opening at the base of the associated fang (Greene 1997). The venom glands of Heloderma are not surrounded by compressor musculature as in most venomous snakes. Instead, tension within the glands produced by jaw movements propels venom toward the venom-conducting teeth, and capillary action carries the venom from grooved teeth into the wound.
The structure of the venom gland was first described by Fischer (1882), but the most complete descriptions to date remain the work of Fox (1913) and Phisalix (1912, 1917). More recent reviews (e.g., Bogert and Martin del Campo 1956; Tinkham 1971b; Russell and Bogert 1981), as well as what I provide below, are largely summaries of this earlier work.
The venom glands of helodermatid lizards are visible externally as conspicuous swellings below the lower lips (fig. 9, plate 20). Each gland is surrounded by a fibrous capsule from which septa extend to divide the gland into three or four distinct lobes (fig. 9). Each lobe of the venom gland is a structurally independent organ that forms a sac with a swollen glandular region at its base and a narrowed excretory duct near the upper end, which empties at the base of the teeth in the lower jaw (fig. 10). Each lobe of the venom gland is subdivided (by septa) into several lobules, which are further subdivided into smaller lobules. These subdivisions continue to occur, resulting in tiny chambers, or alveoli, each separated from one another by a delicate septum. The cavities within the alveoli are continuous with the lobules, which are, in turn, continuous with one another, forming a network of intralobular tubules. It is within these structures, apparently, that venom is produced by columnar, granule-secreting cells that line the intralobular tubules. As these cells discharge their contents, the secreted granules flow from the tubules into a central collecting lumen, which connects to the excretory duct. The lumen, along with associated tubules and alveoli, apparently also serves as a storage reservoir for venom.
Helodermatids are not specialized for injecting large quantities of venom during brief contact, as are many venomous snakes, but the venom delivery system is structured to quickly and effectively transfer venom into a bite. During biting, tension produced in the gland by jaw movements propels venom through the venom ducts into a region between the fourth and seventh pair of dentaries (counting from the front), where the teeth show their greatest specialization for piercing and venom delivery. A series of small folds and grooves in the membranous tissue within this region serves as a temporary reservoir for the venom and may facilitate the flow of venom from duct to tooth. When the lizard bites, venom flows from these reservoirs through the grooved teeth into the wound.
Each specialized tooth has two grooves, with a shallower (sometimes absent) groove toward the rear (fig. 11) . Each groove is flanked by a cutting flange, which makes the tooth better adapted for piercing flesh than a merely conical tooth. Not all the teeth are similar in structure or size. The largest, most deeply grooved teeth are the dentaries (in the lower jaw), which can be up to 6 mm long in H. horridum and 5 mm long in H. suspectum (fig. 11). The maxillary teeth (in the upper jaw) are shorter and less strongly grooved. Some teeth (especially the premaxillaries toward the front of the mouth) are hardly grooved at all, but they can still effectively deposit venom into an adversary (Tinkham 1956; Strimple et al. 1997).
Jaw movements other than those associated with biting may also produce sufficient tension within the glands to bathe all the teeth in venom. An agitated lizard will often display a defensive posture of opening its jaws wide (exposing the purplish-white interior of its mouth) then closing and reopening the jaws as a threat continues. This behavior may serve to bathe the maxillary teeth, dentaries, and premaxillaries in venom preparatory to envenomation.
The quantity of venom deposited into the victim may vary with many factors, including size of the lizard, degree of agitation, and length of time it remains attached. In captive Gila Monsters, quantities ranging from 0.5 to 2.0 ml in a single milking have been extracted by a variety of methods (Loeb et al. 1913; Arrington 1930; Brown and Lowe 1955; Alagon et al. 1982; fig. 12).
EFFECTS OF THE VENOM
The most complete investigation of the effects of Heloderma venom remains the work of Loeb et al. (1913) who tested hundreds of species, including invertebrates. Invertebrates are essentially immune to the effects of Heloderma venom. Effects on vertebrates are more severe and varied. Ectotherms appear markedly less susceptible to the effects of the venom than do endotherms (Cooke and Loeb 1913). Notably, Gila Monsters appear to be immune to the effects of their own venom (Cooke and Loeb 1913; Brown and Lowe 1954).
In mammals, major effects include a rapid reduction in carotid blood flow followed by a marked fall in blood pressure, respiratory irregularities, tachycardia and other cardiac anomalies, as well as hypothermia, edema and internal hemorrhage in the gastrointestinal tract, lungs, eyes, liver, and kidneys (table 4). Common symptoms in dogs and cats include vomiting accompanied by discharge of urine and feces, and copious flow of saliva and tears. Death from large doses of Heloderma venom has been attributed primarily to respiratory disturbances (Cooke and Loeb 1913). Postmortem examinations reveal congestion and edema in the lungs, a marked congestion of the serous layer of the stomach and intestines, and hemorrhage in the kidneys and liver (Cooke and Loeb 1913; Ariano Sanchez 2003). Sublethal doses in mice and rats produce protrusion of the eyes and periorbital bleeding (Cooke and Loeb 1913; Ariano Sanchez 2003), and in rabbits they lead to an increase in the number of white blood cells (Meyers and Tuttle 1913). In humans, the effects of bites are associated with excruciating pain that may extend well beyond the area bitten and persist up to 24 hours. Other common effects of Heloderma bites on humans include local edema (swelling), weakness, sweating, and a rapid fall in blood pressure (see below). A summary of the effects of Heloderma venom on mammals is given in table 4.
The lethal dose ([LD.sub.50]) of Heloderma venom varies among studies and venom lots (Russell and Bogert 1981). Such values are influenced by the relative amounts of venom and saliva collected in each sample and are, therefore, difficult to evaluate. The venom is most toxic when administered intravenously in mice, with [LD.sub.50] values varying from 0.4 to 2.7 mg/kg for H. suspectum (table 5). With an i.v. L[D.sub.50] of 1.4 to 2.7 mg/kg, the venom of H. horridum appears to have toxicity similar to that of H. suspectum. The LD for H. horridum charlesbogerti venom is 1.0 mg/kg when injected intramuscularly in rats (Ariano Sanchez 2003). When injected into mammals, the venom of Heloderma appears to be about as toxic as that of the Western Diamondback Rattlesnake, Crotalus atrox (Russell and Bogert 1981).
CHEMICAL MAKEUP OF THE VENOM
As knowledge of the toxic effects produced by Heloderma venom increased, so did interest in the chemical constituents causing these symptoms. The first studies on the chemical nature of Heloderma venom (Santesson 1897) identified two "toxic principles," referred to as nuclein and albuminose, but it was not until 1913 that the first toxin, a lipase, was isolated (Alsberg 1913). The 1960s saw renewed interest in the chemistry of Heloderma venom. Serotonin and amine oxidase activity were identified in the venom of H. horridum in 1960 (Zarafonetis and Kalas 1960). Mebs and Raudonat (1966) identified a very active hyaluronidase (a spreading factor; see below), phospholipase A, and a kinin-releasing enzyme with small proteolytic activity in both H. horridum and H. suspectum venom. In 1967, Tu and Murdoch showed that H. suspectum venom was largely a mixture of proteins, some of which hydrolyze certain peptides. Patterson and Lee (1969) later showed coagulation was affected if venom was incubated with blood plasma for longer periods. Subsequent work isolated hemorrhagic toxins in the venom (Nikai et al. 1988; see below). In the late 1960s, Mebs isolated kallikrein from the venom of H. suspectum (Mebs 1968, 1969). Murphy et al. (1976) demonstrated enzyme activities in the venom of H. horridum. In the 1980s, additional proteins-gilatoxin (Hendon and Tu 1981), horridum toxin (an arginine ester hydrolase; Alagon et al. 1982; Nikai et al. 1988), helodermatine (a kallikrein-like hypotensive enzyme; Alagon et al. 1986), an additional phospholipase ([A.sub.2]; Gomez et al. 1989), and helothermine (Mochca-Morales et al. 1990)-were discovered and added to the known arsenal of Heloderma venom components.
With the discovery in the 1980s that Heloderma venom contained strongly bioactive agents, with hormonelike actions similar to vasoactive intestinal peptides (VIP), interest and research in these intriguing molecules accelerated dramatically (Raufman 1996). Five bioactive peptides have been isolated so far from the venoms of Gila Monsters and Beaded Lizards: helospectin I and II (Parker et al. 1984), helodermin (Hoshino et al. 1984), exendin-3 (found in H. horridum; Eng et al. 1990), and exendin-4 (found in H. suspectum; Eng et al. 1992). These peptides mimic several human neurosecretory hormones that relax smooth muscle and mediate energy metabolism. The major compounds so far identified in the venom of helodermatid lizards as well as their chemical nature and physiological effects are discussed below and summarized in table 6.
Hyaluronidase is a hydrolase enzyme that cleaves internal glycosidic bonds of hyaluronic acid, a mucopolysaccharide that is an important component of connective tissue. Because this action facilitates venom diffusion into the tissue (Tu and Hendon 1983), hyaluronidase has been termed spreading factor. Hyaluronidase is also common in snake venoms where it also acts as a spreading factor (Meier and Stocker 1995). Venom of both Heloderma species shows particularly high hyaluronidase activity, which is believed to explain the potent edema effects of Heloderma bites (Mebs and Raudonat 1966; Russell and Bogert 1981).
Serotonin, derived from the amino acid tryptophan, produces potent local physiological effects and also functions as a neurotransmitter. Serotonin mediates local processes such as inflammation, vasodilation, and smooth muscle activity and causes aggregation of platelets (Greger 1996). It is found in a variety of toxins from animals such as spiders, toads, and wasps. The specific in vivo effects from the serotonin found in Heloderma venom are not known, although it likely participates in the inflammation response.
Phospholipase [A.sub.2] enzymes are classified as hydrolases that act on ester bonds in fat molecules (phospholipids). These common constituents of viperid and elapid snake venoms are toxic to presynaptic membranes, disrupting the release of neurotransmitters at nerve synapses and neuromuscular junctions and inhibiting platelets (among other actions; Rosenberg 1988; Meier and Stocker 1995). This behavior can cause paralysis and loss of muscle control and function, among other signs. We do not know whether phospholipase [A.sub.2] from Heloderma is responsible for such actions in humans and other mammals because the specific in vivo effects of this enzyme have not been investigated. However, phospholipase [A.sub.2] enzymes from H. horridum have been shown to inhibit platelet aggregation in human blood plasma (Huang and Chiang 1994).
Five variants of phospholipase [A.sub.2] (or [PLA.sub.2]) have been identified and characterized in the venom of H. suspectum (Sosa et al. 1986; Gomez et al. 1989; Vandermeers et al. 1991). All of these stimulate the release of amylase from secretory cells (acini) in rat pancreatic tissue, and they hydrolyze phosphatidylcholines (lecithins), which are important in cell structure and metabolism. Gila Monster [PLA.sub.2]s are apparently quite different from those found in snake venoms, especially in their N-terminal amino acid sequences and molecular weight (Tu 1991; Vandermeers et al. 1991). Interestingly, they are most similar to [PLA.sub.2] from bee venom. The major variant of [PLA.sub.2] in Gila Monsters (Pa5) shows a C-terminal extension seen only in Heloderma and bee (Apis mellifera) venoms (Gomez et al. 1989).
Excerpted from BIOLOGY OF Gila Monsters and Beaded Lizards by DANIEL D. BECK Excerpted by permission.
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