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Phosphodiesterase 1A modulates cystogenesis in zebrafish.

ABSTRACT: Substantial evidence indicates the importance of elevated cAMP in polycystic kidney disease (PKD). Accumulation of cAMP in cystic tissues may be, in part, caused by enhanced adenylyl cyclase activity, but inhibition of cAMP degradation by phosphodiesterases (PDE) likely has an important role, because cAMP is inactivated much faster than it is synthesized. PDE1 is the only PDE family activated by Ca(2+), which is reduced in PKD cells. To assess the contribution of the PDE1A subfamily to renal cyst formation, we examined the expression and function of PDE1A in zebrafish. We identified two splice isoforms with alternative starts corresponding to human PDE1A1 and PDE1A4. Expression of the two isoforms varied in embryos and adult tissues, and both isoforms hydrolyzed cAMP with Ca(2+)/calmodulin dependence. Depletion of PDE1A in zebrafish embryos using splice- and translation-blocking morpholinos (MOs) caused pronephric cysts, hydrocephalus, and body curvature. Human PDE1A RNA and the PKA inhibitors, H89 and Rp-cAMPS, partially rescued phenotypes of pde1a morphants. Additionally, MO depletion of PDE1A aggravated phenotypes in pkd2 morphants, causing more severe body curvature, and human PDE1A RNA partially rescued pkd2 morphant phenotypes, pronephric cysts, hydrocephalus, and body curvature. Together, these data indicate the integral role of PDE1A and cAMP signaling in renal development and cystogenesis, imply that PDE1A activity is altered downstream of polycystin-2, and suggest that PDE1A is a viable drug target for PKD.

SUBMITTER: Sussman CR

PROVIDER: S-EPMC4178429 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature

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Phosphodiesterase 1A modulates cystogenesis in zebrafish.

Sussman Caroline R CR Ward Christopher J CJ Leightner Amanda C AC Smith Jordan L JL Agarwal Reema R Harris Peter C PC Torres Vicente E VE

Journal of the American Society of Nephrology : JASN 20140403 10

Substantial evidence indicates the importance of elevated cAMP in polycystic kidney disease (PKD). Accumulation of cAMP in cystic tissues may be, in part, caused by enhanced adenylyl cyclase activity, but inhibition of cAMP degradation by phosphodiesterases (PDE) likely has an important role, because cAMP is inactivated much faster than it is synthesized. PDE1 is the only PDE family activated by Ca(2+), which is reduced in PKD cells. To assess the contribution of the PDE1A subfamily to renal cys ...[more]

PMID: 24700876

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Project description:Cannabinoid CB1 receptors (CB1Rs) mediate the presynaptic effects of endocannabinoids in the central nervous system (CNS) and most behavioral effects of exogenous cannabinoids. Cannabinoid receptor-interacting protein 1a (CRIP1a) binds to the CB1R C-terminus and can attenuate constitutive CB1R-mediated inhibition of Ca(2+) channel activity. We now demonstrate cellular colocalization of CRIP1a at neuronal elements in the CNS and show that CRIP1a inhibits both constitutive and agonist-stimulated CB1R-mediated guanine nucleotide-binding regulatory protein (G-protein) activity. Stable overexpression of CRIP1a in human embryonic kidney (HEK)-293 cells stably expressing CB1Rs (CB1-HEK), or in N18TG2 cells endogenously expressing CB1Rs, decreased CB1R-mediated G-protein activation (measured by agonist-stimulated [(35)S]GTP?S (guanylyl-5'-[O-thio]-triphosphate) binding) in both cell lines and attenuated inverse agonism by rimonabant in CB1-HEK cells. Conversely, small-interfering RNA-mediated knockdown of CRIP1a in N18TG2 cells enhanced CB1R-mediated G-protein activation. These effects were not attributable to differences in CB1R expression or endocannabinoid tone because CB1R levels did not differ between cell lines varying in CRIP1a expression, and endocannabinoid levels were undetectable (CB1-HEK) or unchanged (N18TG2) by CRIP1a overexpression. In CB1-HEK cells, 4-hour pretreatment with cannabinoid agonists downregulated CB1Rs and desensitized agonist-stimulated [(35)S]GTP?S binding. CRIP1a overexpression attenuated CB1R downregulation without altering CB1R desensitization. Finally, in cultured autaptic hippocampal neurons, CRIP1a overexpression attenuated both depolarization-induced suppression of excitation and inhibition of excitatory synaptic activity induced by exogenous application of cannabinoid but not by adenosine A1 agonists. These results confirm that CRIP1a inhibits constitutive CB1R activity and demonstrate that CRIP1a can also inhibit agonist-stimulated CB1R signaling and downregulation of CB1Rs. Thus, CRIP1a appears to act as a broad negative regulator of CB1R function.

Dataset Information

Phosphodiesterase 1A modulates cystogenesis in zebrafish.

Publications

Phosphodiesterase 1A modulates cystogenesis in zebrafish.

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