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Intracerebroventricular injections of oligonucleotide probes complementary to oxytocin mRNA are known to decrease systemic oxytocin levels. In this study we show that immunoreactive oxytocin in the magnocellular hypothalamic perikarya and in their neurohypophysial projections remains unaffected by intracerebroventricular injections with an oxytocin antisense probe in rats. Hybridization signal for oxytocin mRNA was increased in the supraoptic and paraventricular nuclei in these animals. Immunocytochemistry with a monoclonal antibody, raised against triple helical DNA resulted in an accumulation of cytoplasmic reaction product in many of the magnocellular oxytocin immunoreactive neurons and in a fraction of the Herring bodies inthe posterior pituitary lobe in the antisense treated rats. Such immunostaining could be abolished by pretreating sections with RNase H. Animals injected with a mismatch probe instead of the antisense probe were devoid of cytoplasmic or axonal triple helix immunostaining. Our findings indicate that oxytocinergic transcripts in magnocellular hypothalamic neurons form triple helix-like aggregates upon specific antisense targeting rather than being degraded by endogenous RNases. While de novo transcription of oxytocin is probably stimulated, systemic release of the nonapeptide may be impaired.