Comparison of the neuroprotective effects of adrenoceptor drugs in retinal cell culture and intact retina.
OBJECTIVE
The efficacy of beta1-adrenoceptor (AR)-selective (betaxolol and metoprolol) and nonselective (timolol) antagonists and the alpha2-AR agonist UK14,304 as retinal neuroprotectants was compared and contrasted in an in vitro glutamate excitotoxicity model. The ability of UK14,304, brimonidine, and betaxolol to alter glutamate-receptor-induced changes in intracellular calcium ([Ca2+]i) was also determined in isolated retinal neurons and retinal ganglion cells (RGCs) in an intact retina preparation.


METHODS
Neuronal survival was measured in mixed retinal cell cultures treated for 24 hours with media containing 100 microM glutamate, with or without the addition of each of the drugs (1-1000 microM). Effects of glutamate on glia were also investigated in a C6 glioma cell line. Glutamate-induced changes in [Ca2+]i with and without UK14,304, and its analogue brimonidine were assessed by calcium-imaging techniques in retinal neurons in culture. The effect of betaxolol on [Ca2+]i was investigated in RGCs in intact rabbit retina.


RESULTS
In cell cultures, 10-1000 microM glutamate resulted in a dose-dependent loss of neurons, but not of glia. The absence of glutamate toxicity in glia was confirmed in C6 glioma cells. Betaxolol, but not timolol or metoprolol, significantly increased survival (from 52% of control in glutamate-only to 78% with 10 microM betaxolol) after excitotoxic insult. UK14,304 also increased survival (from 62% of control in glutamate only to 109% and 101% of control with 10 and 100 microM UK14,304, respectively). This effect was blocked by the specific alpha2-antagonist, yohimbine. Both UK14,304 and brimonidine (10-100 microM) reduced glutamate-induced [Ca2+]i increases in retinal neurons in culture. The actions of the alpha2-agonists in reducing glutamate-induced [Ca2+]i increases were reduced by yohimbine (1 microM). Betaxolol (100 microM) reduced N-methyl-D-aspartate (NMDA)-induced increases of [Ca2+]i in RGCs in intact retina.


CONCLUSIONS
Betaxolol reduced glutamate excitotoxicity in retinal neurons in vitro through a mechanism independent of beta-AR interactions. UK14,304, acting through alpha2-ARs, was also neuroprotective in vitro. The neuroprotective actions of betaxolol and the alpha2-agonists on retinal neurons may be due, at least in part, to a direct reduction of glutamate receptor-mediated increases of [Ca2+]i.
Baptiste
Darryl C
DC
Laboratory for Retina and Optic Nerve Research, Dalhousie University, Halifax, Nova Scotia, Canada.
Hartwick
Andrew T E
AT
Jollimore
Christine A B
CA
Baldridge
William H
WH
Chauhan
Balwantray C
BC
Tremblay
François
F
Kelly
Melanie E M
ME
D000316:Adrenergic alpha-Agonists / Q000494:pharmacology*; D000319:Adrenergic beta-Antagonists / Q000494:pharmacology*; D000818:Animals; D015784:Betaxolol / Q000494:pharmacology; D000068438:Brimonidine Tartrate; D002118:Calcium / Q000378:metabolism; D018929:Cell Culture Techniques; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D016257:Fura-2; D018698:Glutamic Acid / Q000494:pharmacology; D008790:Metoprolol / Q000494:pharmacology; D016202:N-Methylaspartate / Q000494:pharmacology; D009457:Neuroglia / Q000187:drug effects / Q000378:metabolism; D018696:Neuroprotective Agents / Q000494:pharmacology*; D011810:Quinoxalines / Q000494:pharmacology; D051381:Rats; D020318:Rats, Long-Evans; D012160:Retina / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D012165:Retinal Ganglion Cells / Q000187:drug effects / Q000378:metabolism; D013999:Timolol / Q000494:pharmacology
D000316:Adrenergic alpha-Agonists; D000319:Adrenergic beta-Antagonists; D018696:Neuroprotective Agents; D011810:Quinoxalines; D018698:Glutamic Acid; D000068438:Brimonidine Tartrate; D016202:N-Methylaspartate; D013999:Timolol; D008790:Metoprolol; D015784:Betaxolol; D002118:Calcium; D016257:Fura-2