Overview of the Issue

Our laboratory has been studying the genetics behind epilepsy and Addison's disease in several dog breeds for a number of years. This report will summarize some of the current findings regarding these two genetic disorders beginning with epilepsy.

DeLahunta (1977) characterizes a seizure as "a transitory disturbance of brain function" representing enhanced and synchronous activity of neurons (March,. 1998). In many instances, the seizure reflects an isolated incident and never recurs. Recurrent seizure activity is what defines epilepsy. Epilepsy is then categorized based upon the cause of the seizures. Seizures can be sequelae to other physiological conditions such as infections, cancer, or hypoglycemia. Seizures can also be the consequence of exposure to environmental toxins or the result of trauma. When a particular initiator for the seizures can be identified, then the epilepsy is referred to as secondary or acquired epilepsy. When no underlying cause can be detected, the epilepsy is considered primary or idiopathic epilepsy. In both categories of epilepsy, the actual neurological disturbance is similar, but the initiator differs.

Idiopathic epilepsy is an extremely common neurological disorder in dogs and cats. The Canine Epilepsy Research Project, a consortium of researchers from the Universities of Missouri and Minnesota, reports having DNA submissions of epileptic dogs from 85 different dog breeds of genetically diverse backgrounds. Epilepsy in cats, while much less prevalent, is still a serious condition (Kline, 1998), although the preponderance of feline seizures represent acquired or secondary epilepsy (Parent and Quesnel, 1996).

Seizures can be mild with symptoms of generalized confusion and "gazing" or seizures can be severe (grand mal) with loss of consciousness, spastic muscle movements, bowel and urinary incontinence, and involuntary salivation. Seizure frequency can vary from several times per day to more intermittent episodes occurring perhaps once every few years (Thomas, 2000). Although the duration of a seizure is generally short, the after effects may linger for hours to days following the episode itself. Further, the occurrence of one seizure may potentiate future seizures (March, 1998). Regardless of precisely how the seizures are manifested, the effects of idiopathic epilepsy are distressing for the dog and the owner, and possibly even hazardous in the larger breeds.

Many diverse dog breeds experience seizures and while the majority of breeds are considered to have a genetic component to the epilepsy, the details of inheritance in most breeds are not well described. For any sound inheritance study, the first step to characterizing the genetic contribution is to obtain sufficient phenotypic and pedigree data from related dogs. One significant complication to the study of epilepsy is the ambiguity of diagnosis. Idiopathic epilepsy diagnosis is one of exclusion in that no underlying cause for the seizures can be identified. Therefore, it is vital that the families studied transmit very definable, recognizable seizure activity through the generations. Further, any study needs to account that age of seizure onset, while typically between 2-4 years of age, can vary so designating a dog as "unaffected" needs to be done judiciously.

In our studies we have restricted our analyses to dogs that exhibit grand mal seizuring on the premise that owners can readily identify that seizure form. We are presently studying epilepsy in the Belgian Tervuren, Belgian Sheepdogs, Poodles, English Mastiffs, and Giant Schnauzers. By far we have made the most progress on the Belgian breeds as we have been studying those for the greatest length of time. However, the approach our laboratory has taken with the Belgians is the same as for the other breeds. We have collected phenotypic (health status) and pedigree data, along with DNA samples, on more than 1850 Belgians (with an average of 12.75% classified as epileptic). These samples complement an earlier "phenotype-only" study of ~ 1000 Belgians. From these data we have determined, statistically, that the heritability for seizures in the Belgians is on the order of 0.77 to 0.83 (Famula et al., 1997; Famula and Oberbauer, 1998) indicating a large genetic component to the expression of seizures. As an aside, the heritability estimate for epilepsy in the English Mastiff is likewise, extremely high. The data for the Belgians were then subjected to complex segregation analyses to characterize the mode of inheritance which suggested, polygenic with a single major gene of large effect inherited as an autosomal recessive influencing the expression of the disorder.

These findings formed the rationale in undertaking a full scale genome scan of the Belgian DNA to identify a genetic region linked to the seizure phenotype. We initially scanned a small cohort of related dogs with microsatellite markers offering reasonable genome coverage (Oberbauer et al., 2003). We have since expanded the number of dogs evaluated as well as the extent of genomic coverage. Five genomic regions have been identified as potentially linked to the expression with one region exhibiting fairly robust LOD scores. LOD scores are defined as the log (base 10) of a likelihood ratio between two conditional probabilities, one being that linkage exists and the other being that the marker and phenotype are unlinked. In human studies, LOD scores in excess of 3.0 are considered "significant" for linkage (Shete and Amos, 2002). One genomic region has consistently yielded LOD scores hovering around 3.0. We are currently expanding the number of dogs analyzed and the number of genetic markers with the intent to enhance the LOD scores and improve the confidence of linkage. With linkage established, the DNA will be sequenced and candidate genes within that region will be investigated for causal mutations. The ultimate objective is to generate a genetic marker test to allow breeders to identify dogs that carry a mutation permissive for the expression of epilepsy.

Similar heritability and complex segregation studies have been ongoing in other breeds (e.g., Patterson et al., 2005). In January of this year, a research team in Canada uncovered the causal mutation for a very specific form of epilepsy (progressive myoclonic epilepsy inherited as an autosomal recessive) that plagues miniature wirehaired dachshunds (Lohi et al., 2005). The genomic scanning approach resulted in the development of a genetic test that is now available for breeders of the miniature wirehaired dachshund.

The current research on the genetics of Addison's disease, or hypoadrenocorticism as it is more accurately termed, also shows a very strong genetic contribution to the expression of the disorder. In Addison's disease, the adrenal cortex fails to synthesize and release adequate quantities of two classes of steroid hormones. Mineralocorticoids that regulate electrolyte balance and corticosteroids that regulate many aspects of metabolism and the stress response are greatly reduced in dogs with Addison's.

Similar to epilepsy, Addison's disease can be characterized as being either primary or secondary. As the name implies, primary reflects an insufficiency due to a defect or atrophy in the adrenal gland itself. Secondary Addison's disease reflects the condition where the impaired adrenal cortex function is the consequence of some other identifiable cause; for example, a deficiency in adrenocorticotropin hormone (ACTH), the hormone that stimulates the adrenal gland to function.

As noted above, with primary Addison's, the adrenal cortex gradually deteriorates and becomes incapable of hormonal production (Kaufman, 1984); this deterioration is speculated to be a consequence of the immune system failing to distinguish that the adrenal cortex is a self tissue (Smallwood and Barsanti, 1995; Greco and Harpold, 1994; Weller et al., 1996). Thus, Addison's disease is often a late onset disorder with diffuse symptoms that include generalized fatigue, inappetence, gastrointestinal upset, and weight loss. Primary Addison's seems to be the most prevalent form and is diagnosed by providing exogenous ACTH and evaluating the adrenal cortex's ability to secrete glucocorticoids. If ACTH fails to induce glucocorticoid production and release, the adrenal cortex is considered defective and the animal is determined to be a primary Addisonian.

The existence of Addisonian dogs repeatedly appearing in pedigrees of certain dogs led breeders to speculate that Addison's disease is inherited. Although Addison's disease occurs in the dog population as a whole (Little et al., 1989), within certain breeds there has been a higher than expected incidence noted. We have determined the heritability and mode of inheritance if feasible in the Standard Poodle, Great Dane, West Highland White Terrier, Bearded Collie, Portuguese Water Dog, and Leonberger. Using an approach identical to that described for the epilepsy work, we have estimated heritability for those breeds with sufficient data submission. In all breeds except the Great Dane (which currently lacks the necessary numbers of dogs enrolled in the study) the heritability for Addison's disease is greater than 0.7 indicating a very large degree of genetic regulation.

Complex segregation analyses confirm the genetic component and suggest that the best fit mode of inheritance is autosomal recessive with modifying genes. In other words, Addison's appears to be polygenic but with a major controlling gene. The lesser genes likely regulate the age of onset and the progression of the disorder. Of note, is that there is no sex affect in any of the breeds reflecting an equal number of males and females diagnosed with Addison's disease.

Based upon these findings, we have approached the genetic linkage study in three separate ways. 1) Using multigenerational families of Poodles and Portuguese Water Dogs, we are scanning for linkage between the disorder phenotype and genetic markers linked to candidate loci. The candidate genes were chosen for their involvement in normal immune function and cell recognition. At this point in time, no linkage among the genes tested was discerned. 2) For Poodles, Bearded Collies, and Portuguese Water Dogs, we are using eight highly unrelated dogs of each breed in a homozygosity screen using 327 microsatellite markers that offer 9 MB genome coverage. The theory is that since the major gene is autosomal recessive, the unrelated dogs should show homozygosity in the chromosomal region that regulates the expression but not in other chromosomal regions. This is because, on the whole, the dogs chosen are not closely related and therefore should share little DNA similarity other than in that one region (Lohi et al., 2005). Several chromosomal regions indicate further investigation is warranted and we will target those regions with additional markers. 3) Using multigenerational families of the above breeds we will use the 327 markers in a full genome scan. We believe this approach will yield chromosomal regions significantly linked to the Addisonian phenotype. The linkage, followed by gene identification and genetic sequencing to identify the precise mutation will enable the development of a diagnostic DNA based test for breeders to integrate into their breeding program. We are hopeful that the diversity of breeds under study will result in a test that is of utility to all breeds.

References

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8.  Parent JM, Quesnel AD. 1996. Seizures in cats. Vet Clin North Am Small Anim Pract. 26:811-25.

9.  Patterson EE, Armstrong PJ, O'Brien DP, Roberts MC, Johnson GS, Mickelson JR. 2005. Clinical description and mode of inheritance of idiopathic epilepsy in English springer spaniels. J Am Vet Med Assoc. 226:54-8.

10. Shete S, Amos CI. 2002. Testing for genetic linkage in families by a variance components approach in the presence of genomic imprinting. Am J Hum Genet 70:751 7

11. Smallwood LJ, Barsanti JA. 1995. Hypoadrenocorticism in a family of leonbergers. J Am Anim Hosp Assoc. 31:301-5.

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13. Weller RE, Buschbom RL, Dagle GE, Park JF, Ragan HA, Watson CR. 1996. Hypoadrenocorticism in beagles exposed to aerosols of plutonium-238 dioxide by inhalation. Radiat Res. 146:688-93.