␣1 subunit of the voltage-dependent Ca 2؉ channel is essential for channel function and determines the functional specificity of various channel types. ␣1E subunit was originally identified as a neuron-specific one, but the physiological function of the Ca 2؉ channel containing this subunit (␣1E Ca 2؉ channel) was not clear compared with other types of Ca 2؉ channels because of the limited availability of specific blockers. To clarify the physiological roles of the ␣1E Ca 2؉ channel, we have generated ␣1E mutant (␣1E؊͞؊) mice by gene targeting. The lacZ gene was inserted in-frame and used as a marker for ␣1E subunit expression. ␣1E؊͞؊ mice showed reduced spontaneous locomotor activities and signs of timidness, but other general behaviors were apparently normal. As involvement of ␣1E in pain transmission was suggested by localization analyses with 5-bromo-4-chloro-3-indolyl ␤-D-galactopyranoside staining, we conducted several pain-related behavioral tests using the mutant mice. Although ␣1E؉͞؊ and ␣1E؊͞؊ mice exhibited normal pain behaviors against acute mechanical, thermal, and chemical stimuli, they both showed reduced responses to somatic inflammatory pain. ␣1E؉͞؊ mice showed reduced response to visceral inflammatory pain, whereas ␣1E؊͞؊ mice showed apparently normal response compared with that of wild-type mice. Furthermore, ␣1E؊͞؊ mice that had been presensitized with a visceral noxious conditioning stimulus showed increased responses to a somatic inflammatory pain, in marked contrast with the wild-type mice in which long-lasting effects of descending antinociceptive pathway were predominant. These results suggest that the ␣1E Ca 2 ؉ channel controls pain behaviors by both spinal and supraspinal mechanisms.V oltage-dependent calcium channels (VDCCs) are classified into several distinct groups termed L-, N-, P-, Q-, R-, and T-types (1, 2). These types of VDCCs play important roles in various neuronal activities, including the control of neurotransmitter release, membrane excitability, and gene expression (3), but exact roles of each channel type are not necessarily clarified. In particular, functions of the R-type Ca 2ϩ channel are least understood. The R-type Ca 2ϩ channel was originally defined as a channel ''Resistant'' to blockers for L-, N-, P-, and Q-type Ca 2ϩ channels (4); therefore, it is possible that the R-type current is a mixture of several different drug-resistant Ca 2ϩ currents. Although the R-type Ca 2ϩ channel is suggested to play a critical role in the release of neurotransmitters and somatodendritic excitability in a certain set of neurons (4-6), the physiological functions of this channel remain to be clarified.VDCCs are heteromultimers composed of ␣ 1 , ␣ 2 -␦, ␤, and ␥ subunits. ␣ 1 subunit is essential for channel function and determines the type of each Ca 2ϩ channel. So far, 10 different ␣ 1 cDNAs (␣ 1A-I and ␣ 1S ) have been cloned from a variety of tissues, and extensive studies have been made to clarify the relationship between each cloned ␣ 1 subunit and native Ca 2ϩ channels (2)....

The brain stores and recalls memories through a set of neurons, termed engram cells. However, it is unclear how these cells are organized to constitute a corresponding memory trace. We established a unique imaging system that combines Ca 2+ imaging and engram identification to extract the characteristics of engram activity by visualizing and discriminating between engram and non-engram cells. Here, we show that engram cells detected in the hippocampus display higher repetitive activity than non-engram cells during novel context learning. The total activity pattern of the engram cells during learning is stable across post-learning memory processing. Within a single engram population, we detected several sub-ensembles composed of neurons collectively activated during learning. Some sub-ensembles preferentially reappear during post-learning sleep, and these replayed sub-ensembles are more likely to be reactivated during retrieval. These results indicate that sub-ensembles represent distinct pieces of information, which are then orchestrated to constitute an entire memory.

The extent to which Ca2+-induced Ca2+ release (CICR) affects transmitter release is unknown. Continuous nerve stimulation (20–50 Hz) caused slow transient increases in miniature end-plate potential (MEPP) frequency (MEPP-hump) and intracellular free Ca2+ ([Ca2+]i) in presynaptic terminals (Ca2+-hump) in frog skeletal muscles over a period of minutes in a low Ca2+, high Mg2+ solution. Mn2+ quenched Indo-1 and Fura-2 fluorescence, thus indicating that stimulation was accompanied by opening of voltage-dependent Ca2+ channels. MEPP-hump depended on extracellular Ca2+ (0.05–0.2 mM) and stimulation frequency. Both the Ca2+- and MEPP-humps were blocked by 8-(N,N-diethylamino)octyl3,4,5-trimethoxybenzoate hydrochloride (TMB-8), ryanodine, and thapsigargin, but enhanced by CN−. Thus, Ca2+-hump is generated by the activation of CICR via ryanodine receptors by Ca2+ entry, producing MEPP-hump. A short interruption of tetanus (<1 min) during MEPP-hump quickly reduced MEPP frequency to a level attained under the effect of TMB-8 or thapsigargin, while resuming tetanus swiftly raised MEPP frequency to the previous or higher level. Thus, the steady/equilibrium condition balancing CICR and Ca2+ clearance occurs in nerve terminals with slow changes toward a greater activation of CICR (priming) during the rising phase of MEPP-hump and toward a smaller activation during the decay phase. A short pause applied after the end of MEPP- or Ca2+-hump affected little MEPP frequency or [Ca2+]i, but caused a quick increase (faster than MEPP- or Ca2+-hump) after the pause, whose magnitude increased with an increase in pause duration (<1 min), suggesting that Ca2+ entry-dependent inactivation, but not depriming process, explains the decay of the humps. The depriming process was seen by giving a much longer pause (>1 min). Thus, ryanodine receptors in frog motor nerve terminals are endowed with Ca2+ entry-dependent slow priming and fast inactivation mechanisms, as well as Ca2+ entry-dependent activation, and involved in asynchronous exocytosis. Physiological significance of CICR in presynaptic terminals was discussed.