OBJECTIVE Phosphodiesterase 4 (PDE4), specific for cyclic AMP (cAMP)-hydrolyzing, has four isoforms (PDE4A-D) with at least 25 splice variants.PDE4 inhibitors produce definite antidepressant-like and cognitive-enhancing effects.However, none of PDE4 inhibitors has yet been approved for clinical utility so far due to the concomitant side effects which appear to be PDE4 isoform specific.The present research is to explore the major isoforms of PDE4 responsible for antidepressant-like and cognitive-enhancing effects of PDF4 inhibitors but not side effects.METHODS Long-form PDE4Ds were silenced by the bilateral microinfusion of lentiviral vector containing miRNAs (4DmiR) into the prefrontal cortex (PFC).The expression of the miRNAs, which were indicated by EGFP, was identified using fluorescence microscopy.Antidepressant-like behaviors were measured by tail-suspension test (TST), forced swimming test (FST) and chronic unpredictable stress model.Learning and memory behaviors were measured by novel object recognition test (NOR) and Morris water maze test (MWM) in both normal mice and the mice with chronic unpredictable stress-induced memory deficits.The emetic potential was evaluated by the assessment of the anaesthetic reversal effect, a surrogate of the emesis test in non-vomiting species.The expressions of PDE4 isoforms/splice variants and cAMP signaling related proteins in PFC were examined by Western-blot analysis.RESULTS (1) High and specific expression of EGFP (green) in PFC was observed under fluorescence microscopy.(2) 4DmiR significantly down-regulated PDE4D4/5 splice variants, but not PDE4A, PDE4B or PDE4D1/2/3.(3) 4DmiR treatments significantly increased cAMP, pCREB and BDNF in PFC.(4) Rolipram and/or 4DmiR treatments significantly decreased immobility in TST and FST.(5) Rolipram and/or 4DmiR treatments reversed the depressive-like behaviors in chronically stressed mice, including the reduced sucrose preference, prolonged latency to novelty-suppressed feeding and increased immobility in FST.(6) Rolipram and/or 4DmiR treatments significantly increased the recognition index in NOR task and both the entries and durations in MWM task.(7) Rolipram and/or 4DmiR treatments reversed the memory deficits in chronically stressed mice, including the reduced the recognition index in NOR task and the decreased durations in MWM task.(8) Rolipram or plus 4DmiR treatment significantly decreased the duration of anaesthesia in the alpha2 adrenergic receptor-mediated anesthesia, but not 4DmiR treatment alone.CONCLUSION Long-form PDE4Ds, at least PDE4D4 and PDE4DS, are the major isoforms responsible for antidepressant-like and cognitive-enhancing effects of PDE4 inhibitors with minor side effects.The critical roles of long-form PDE4Ds in depression and memory are attributed to their regulation of cAMP signaling pathway.