Importance of Temperature in Slice Preparation

1) Cold slice preparation appears to be the cause of the excess spinogenesis. See the following publications that track this phenomenon and how we resolved it:

  • Kirov SA, Harris KM (1999) Dendrites are more spiny on mature hippocampal neurons when synapses are inactivated. Nature Neuroscience 2(10):878-883. 
  • Kirov SA, Sorra KE, Harris KM (1999) Slices have more synapses than perfusion-fixed hippocampus from both young and mature rats. J Neuroscience 19(8):2876-2886. 
  • Fiala JC, Kirov SA, Feinberg MD, Petrak LJ, Goddard CA, George P and Harris KM (2003) Timing of Neuronal and Glial Ultrastructure Disruption During Brain Slice Preparation and Recovery During Incubation In Vitro. J Comp Neurol. 465(1):90-103. 
  • Kirov SA, Petrak LJ, Fiala JC, Harris KM (2004) Dendritic spines disappear with chilling but proliferate excessively upon rewarming of mature hippocampus. Neuroscience 127(1):69-80. (Fig. 9 is especially revealing.)
  • Bourne JN, Kirov SA, Sorra KE, and Harris KM (2007) Warmer preparation of hippocampal slices prevents synapse proliferation that might obscure LTP-related structural plasticity. Neuropharmacol 52:55-59. (See Fig. 2.)

2) Using warm slice preparations with fast chopping (<5 minutes from decapitation until slice in recovery chamber) then need 3 hours of slice recovery plus >1 hour of test pulses to return spine density to perfusion-fixed levels:

  • Bourne JN, Harris KM (2011) Coordination of size and number of excitatory and inhibitory synapses results in a balanced structural plasticity along mature hippocampal CA1 dendrites during LTP. Hippocampus, 21(4):354-73. (Figures 5D, E control - replotted for clarity in Bell et al., listed next.)
  • Bell ME, Bourne JN, Chirillo MA, Mendenhall JM, Kuwajima M, Harris KM (2014) Dynamics of nascent and active zone ultrastructure as synapses enlarge during long-term potentiation in mature hippocampus. J Comp Neurol, 522(17):3861-84. (See Fig. 6.)