Next, to test whether 5-HT acts directly on granule cells to modulate their excitability, we performed current-clamp recordings in the presence of synaptic transmission blockers (NBQX 5 M, R-CPP 5 M, SR95531 5 M) (Fig

Next, to test whether 5-HT acts directly on granule cells to modulate their excitability, we performed current-clamp recordings in the presence of synaptic transmission blockers (NBQX 5 M, R-CPP 5 M, SR95531 5 M) (Fig. 5-HT significantly increases tonic inhibition onto both granule cells and Golgi cells. 5-HT-mediated Golgi cell depolarization is not sufficient, however, to alter the probability or timing of mossy fiber-evoked feed-forward inhibition onto granule cells. Together, increased granule cell tonic inhibition paired with normal feed-forward inhibition acts to reduce granule cell spike probability without altering spike timing. Hence, these data provide a circuit mechanism by which 5-HT can reduce granule cell activity without altering temporal representations of mossy fiber input. Such changes in network integration could enable flexible, state-specific suppression of cerebellar sensorimotor input that should not be learned or enable reversal learning for unwanted associations. NEW & NOTEWORTHY Serotonin (5-hydroxytryptamine, 5-HT) regulates synaptic integration at the input stage of cerebellar processing by increasing tonic inhibition of granule cells. This circuit mechanism reduces the probability of granule cell spiking without altering spike timing, thus suppressing cerebellar input without altering its temporal representation in the granule cell layer. male Sprague-Dawley rats. Slices were prepared in an ice-cold solution of 130 mM K-gluconate, 15 mM KCl, 0.05 mM EGTA, 20 mM HEPES, and 25 mM glucose (pH 7.4), with 2.5 MRS1477 M R-CPP. This solution has previously been found to enhance the survival and health of cerebellar Golgi cells (Hull and Regehr 2012; MRS1477 Kanichay and Silver 2008). Slices were then stored in artificial cerebrospinal fluid containing (in mM) 125 NaCl, 26 NaHCO3, 1.25 NaH2PO4, 2.5 KCl, 1 MgCl2, 2 CaCl2, and 25 glucose and equilibrated with 95% O2 and 5% CO2. This solution contains divalent cation concentrations (3 mM) that permit Golgi cell spontaneous pacemaking in acute rat cerebellar slices. Slices were incubated at 34C for 20 min after preparation, and then kept at room temperature for up to 6 h. Slices were viewed using Dodt Gradient Contrast optics (Scientifica) on an upright microscope (Olympus BX51WI), with a 40 water-immersion objective and a CMOS camera (QImaging, Rolera Bolt). Whole cell and cell-attached recordings were obtained with patch pipettes [Golgi cells: 3C5 M, granule cells, whole cell: 6C9 M, granule cells, cell-attached: 10C14 (M) pulled from borosilicate capillary glass (World Precision Devices) having a Sutter P-1000 micropipette puller]. Electrophysiological recordings were performed at 31C33C. Spontaneous inhibitory postsynaptic currents (sIPSCs) were recorded at 0 mV. Evoked excitatory postsynaptic currents (eEPSCs) were recorded at ?70 mV. The reversal potential for evoked inhibitory postsynaptic currents (eIPSCs) in granule cells was identified empirically in each experiment by modifying the membrane potential (and ?and4 0.05, two asterisks representing 0.01, and three asterisks representing 0.001. Open in a separate windows Fig. 2. 5-Hydroxytryptamine (5-HT) depolarizes Golgi cells by activating 5-HT2A receptors. = 10 Golgi cells treated with 5-HT, no MDL present) and in MDL (gray circles, = 4 Golgi cells treated with 5-HT, MDL present). = 7) and MDL (gray circles, = 4). = 6) and MDL (gray, = 5) as measured by a 5 mV test pulse. * 0.05. Open in a separate windows Fig. 4. 5-Hydroxytryptamine (5-HT) raises spontaneous inhibition and tonic holding current on granule cells inside a 5-HT2AR-dependent manner. = 13 granule cells treated MRS1477 with 5-HT, no MDL or gabazine present) and MDL (gray circles, = Rabbit Polyclonal to TBX2 5 granule cells treated with 5-HT, MDL present; = 8 granule cells treated with 5-HT, gabazine present) (= 9) and MDL (gray circles, = 7) ( 0.05. RESULTS To test whether 5-HT functions presynaptically to modulate excitatory input from mossy materials entering the granule cell coating or postsynaptically to modulate either of the two principal cell.