Rodent hippocampal slices of 0.5 mm thickness have been widely used as a convenient in vitro model since the 1970s. However, spontaneous population rhythmic activities do not consistently occur in this preparation due to limited network connectivity. To overcome this limitation, we develop a novel slice preparation of 1 mm thickness from adult mouse hippocampus by separating dentate gyrus from CA3/CA1 areas but preserving dentate-CA3-CA1 connectivity. While superfused in vitro at 32°C or 37°C, the thick slice exhibits robust spontaneous network rhythms of 1-4 Hz that originate from the CA3 area. Via assessing tissue O₂, K⁺, pH, synaptic and single-cell activities of superfused thick slices, we verify that these spontaneous rhythms are not a consequence of hypoxia and non-specific experimental artifacts. We suggest that the thick slice contains a unitary circuitry sufficient to generate intrinsic hippocampal network rhythms and this preparation is suitable for exploring the fundamental properties and plasticity of a functionally defined hippocampal "lamella" in vitro.