Reduced levels of dopamine and altered metabolism in brains of HPRT knock-out rats: a new rodent model of Lesch-Nyhan Disease

Lesch-Nyhan disease (LND) is a severe neurological disorder caused by loss-of-function mutations in the gene encoding hypoxanthine phosphoribosyltransferase (HPRT), an enzyme required for efficient recycling of purine nucleotides. Although this biochemical defect reconfigures purine metabolism and leads to elevated levels of the breakdown product urea, it remains unclear exactly how loss of HPRT activity disrupts brain function. As the rat is the preferred rodent experimental model for studying neurobiology and diseases of the brain, we used genetically-modified embryonic stem cells to generate an HPRT knock-out rat. Male HPRT-deficient rats were viable, fertile and displayed normal caged behaviour. However, metabolomic analysis revealed changes in brain biochemistry consistent with disruption of purine recycling and nucleotide metabolism. Broader changes in brain biochemistry were also indicated by increased levels of the core metabolite citrate and reduced levels of lipids and fatty acids. Targeted MS/MS analysis identified reduced levels of dopamine in the brains of HPRT-deficient animals, consistent with deficits noted previously in human LND patients and HPRT knock-out mice. The HPRT-deficient rat therefore provides a new experimental platform for future investigation of how HPRT activity and disruption of purine metabolism affects neural function and behaviour

Attenuated variants of Lesch-Nyhan disease

Lesch–Nyhan disease is a neurogenetic disorder caused by deficiency of the enzyme hypoxanthine–guanine phosphoribosyltransferase. The classic form of the disease is described by a characteristic syndrome that includes overproduction of uric acid, severe generalized dystonia, cognitive disability and self-injurious behaviour. In addition to the classic disease, variant forms of the disease occur wherein some clinical features are absent or unusually mild. The current studies provide the results of a prospective and multi-centre international study focusing on neurological manifestations of the largest cohort of Lesch–Nyhan disease variants evaluated to date, with 46 patients from 3 to 65 years of age coming from 34 families. All had evidence for overproduction of uric acid. Motor abnormalities were evident in 42 (91%), ranging from subtle clumsiness to severely disabling generalized dystonia. Cognitive function was affected in 31 (67%) but it was never severe. Though none exhibited self-injurious behaviours, many exhibited behaviours that were maladaptive. Only three patients had no evidence of neurological dysfunction. Our results were compared with a comprehensive review of 78 prior reports describing a total of 127 Lesch–Nyhan disease variants. Together these results define the spectrum of clinical features associated with hypoxanthine–guanine phosphoribosyltransferase deficiency. At one end of the spectrum are patients with classic Lesch–Nyhan disease and the full clinical phenotype. At the other end of the spectrum are patients with overproduction of uric acid but no apparent neurological or behavioural deficits. Inbetween are patients with varying degrees of motor, cognitive, or behavioural abnormalities. Recognition of this spectrum is valuable for understanding the pathogenesis and diagnosis of all forms of hypoxanthine–guanine phosphoribosyltransferase deficiency

Hypoxanthine-guanine phosophoribosyltransferase (HPRT) deficiency: Lesch-Nyhan syndrome

Deficiency of hypoxanthine-guanine phosphoribosyltransferase (HPRT) activity is an inborn error of purine metabolism associated with uric acid overproduction and a continuum spectrum of neurological manifestations depending on the degree of the enzymatic deficiency. The prevalence is estimated at 1/380,000 live births in Canada, and 1/235,000 live births in Spain. Uric acid overproduction is present inall HPRT-deficient patients and is associated with lithiasis and gout. Neurological manifestations include severe action dystonia, choreoathetosis, ballismus, cognitive and attention deficit, and self-injurious behaviour. The most severe forms are known as Lesch-Nyhan syndrome (patients are normal at birth and diagnosis can be accomplished when psychomotor delay becomes apparent). Partial HPRT-deficient patients present these symptoms with a different intensity, and in the least severe forms symptoms may be unapparent. Megaloblastic anaemia is also associated with the disease. Inheritance of HPRT deficiency is X-linked recessive, thus males are generally affected and heterozygous female are carriers (usually asymptomatic). Human HPRT is encoded by a single structural gene on the long arm of the X chromosome at Xq26. To date, more than 300 disease-associated mutations in the HPRT1 gene have been identified. The diagnosis is based on clinical and biochemical findings (hyperuricemia and hyperuricosuria associated with psychomotor delay), and enzymatic (HPRT activity determination in haemolysate, intact erythrocytes or fibroblasts) and molecular tests. Molecular diagnosis allows faster and more accurate carrier and prenatal diagnosis. Prenatal diagnosis can be performed with amniotic cells obtained by amniocentesis at about 15–18 weeks' gestation, or chorionic villus cells obtained at about 10–12 weeks' gestation. Uric acid overproduction can be managed by allopurinol treatment. Doses must be carefully adjusted to avoid xanthine lithiasis. The lack of precise understanding of the neurological dysfunction has precluded development of useful therapies. Spasticity, when present, and dystonia can be managed with benzodiazepines and gamma-aminobutyric acid inhibitors such as baclofen. Physical rehabilitation, including management of dysarthria and dysphagia, special devices to enable hand control, appropriate walking aids, and a programme of posture management to prevent deformities are recommended. Self-injurious behaviour must be managed by a combination of physical restraints, behavioural and pharmaceutical treatments

Purine nucleosides protect injured neurons and stimulate neuronal regeneration by intracellular and membrane receptor-mediated mechanisms

Like adenine-based purines, extracellular nonadenine-based purines have a multitude of trophic effects on the growth, differentiation, and survival of target cells. The nonadenine-based purines, which include guanosine, inosine, and GTP, apparently exert their trophic effects by interacting with both intercellular targets as well as those on the cell surface. Specifically, guanosine and inosine target the protein kinase N-kinase, in promoting remarkable nerve process extension, even in long tracts of the central nervous system after injury. In contrast, GTP may exert its effects via a cell surface receptor coupled to the release of calcium from internal stores. In other cases trophic effects may be mediated by the enhancement of release of adenine-based purines by guanosine. Additionally, evidence is presented for the existence of a high-affinity binding site for guanosine with receptor-like characteristics on the plasma membranes of astrocytes and brain tissue. This site may be G-protein-coupled and exert its effects through activation of the MAP kinase cascade. One effect apparently mediated through this mechanism is the production and release by astrocytes of trophic protein growth factors such as NGF and TGF\u3b2. These have substantial neuroprotective effects. Additionally, this pathway is apparently involved in modulating the expression of P2Y1 and P2Y2 receptors in response to extracellular guanosine. Extracellular nonadenine-based purines can interact with other growth factors, but these interactions are not always synergistic. For example, combinations of guanosine and FGF are antagonistic and reduce the growth of microvascular cells in vitro. Some of the properties of the nonadenine-based purines likely derive from their unique intracellular metabolism in which conversion of guanine to xanthine is the final catabolic step. This step is catalyzed by guanase, the activity of which varies markedly in different brain regions, raising the possibility that guanine or guanosine are involved in neurotransmission. Together these data suggest several potentially useful pharmacological approaches involving nonadenine-based purines to modulate trophic effects in the central nervous syste

Purine nucleosides protect injured neurons and stimulate neuronal regeneration by intracellular and membrane receptor-mediated mechanisms

Like adenine-based purines, extracellular nonadenine-based purines have a multitude of trophic effects on the growth, differentiation, and survival of target cells. The nonadenine-based purines, which include guanosine, inosine, and GTP, apparently exert their trophic effects by interacting with both intercellular targets as well as those on the cell surface. Specifically, guanosine and inosine target the protein kinase N-kinase, in promoting remarkable nerve process extension, even in long tracts of the central nervous system after injury. In contrast, GTP may exert its effects via a cell surface receptor coupled to the release of calcium from internal stores. In other cases trophic effects may be mediated by the enhancement of release of adenine-based purines by guanosine. Additionally, evidence is presented for the existence of a high-affinity binding site for guanosine with receptor-like characteristics on the plasma membranes of astrocytes and brain tissue. This site may be G-protein-coupled and exert its effects through activation of the MAP kinase cascade. One effect apparently mediated through this mechanism is the production and release by astrocytes of trophic protein growth factors such as NGF and TGFβ. These have substantial neuroprotective effects. Additionally, this pathway is apparently involved in modulating the expression of P2Y1 and P2Y2 receptors in response to extracellular guanosine. Extracellular nonadenine-based purines can interact with other growth factors, but these interactions are not always synergistic. For example, combinations of guanosine and FGF are antagonistic and reduce the growth of microvascular cells in vitro. Some of the properties of the nonadenine-based purines likely derive from their unique intracellular metabolism in which conversion of guanine to xanthine is the final catabolic step. This step is catalyzed by guanase, the activity of which varies markedly in different brain regions, raising the possibility that guanine or guanosine are involved in neurotransmission. Together these data suggest several potentially useful pharmacological approaches involving nonadenine-based purines to modulate trophic effects in the central nervous syste